Peve molecular energetic state gene expression analysis ================ 2025-02-19 Analysis of gene expression of P evermanni energetic state using the following gene set GO categories: - Glycolysis - Gluconeogenesis - Lipolysis/lipid catabolism - Fatty acid beta oxidation - Starvation - Lipid biosynthesis - Protein catabolic process # Set up Load libraries ``` r library(ggplot2) library(vegan) ``` ## Loading required package: permute ## Loading required package: lattice ## This is vegan 2.6-6.1 ``` r library(mixOmics) ``` ## Loading required package: MASS ## ## Loaded mixOmics 6.28.0 ## Thank you for using mixOmics! ## Tutorials: http://mixomics.org ## Bookdown vignette: https://mixomicsteam.github.io/Bookdown ## Questions, issues: Follow the prompts at http://mixomics.org/contact-us ## Cite us: citation('mixOmics') ``` r library(readxl) library(factoextra) ``` ## Welcome! Want to learn more? See two factoextra-related books at https://goo.gl/ve3WBa ``` r library(ggfortify) library(ComplexHeatmap) ``` ## Loading required package: grid ## ======================================== ## ComplexHeatmap version 2.20.0 ## Bioconductor page: http://bioconductor.org/packages/ComplexHeatmap/ ## Github page: https://github.com/jokergoo/ComplexHeatmap ## Documentation: http://jokergoo.github.io/ComplexHeatmap-reference ## ## If you use it in published research, please cite either one: ## - Gu, Z. Complex Heatmap Visualization. iMeta 2022. ## - Gu, Z. Complex heatmaps reveal patterns and correlations in multidimensional ## genomic data. Bioinformatics 2016. ## ## ## The new InteractiveComplexHeatmap package can directly export static ## complex heatmaps into an interactive Shiny app with zero effort. Have a try! ## ## This message can be suppressed by: ## suppressPackageStartupMessages(library(ComplexHeatmap)) ## ======================================== ``` r library(viridis) ``` ## Loading required package: viridisLite ``` r library(lme4) ``` ## Loading required package: Matrix ``` r library(lmerTest) ``` ## ## Attaching package: 'lmerTest' ## The following object is masked from 'package:lme4': ## ## lmer ## The following object is masked from 'package:stats': ## ## step ``` r library(emmeans) ``` ## Welcome to emmeans. ## Caution: You lose important information if you filter this package's results. ## See '? untidy' ``` r library(broom.mixed) library(broom) library(tidyverse) ``` ## ── Attaching core tidyverse packages ──────────────────────── tidyverse 2.0.0 ── ## ✔ dplyr 1.1.4 ✔ readr 2.1.5 ## ✔ forcats 1.0.0 ✔ stringr 1.5.1 ## ✔ lubridate 1.9.3 ✔ tibble 3.2.1 ## ✔ purrr 1.0.2 ✔ tidyr 1.3.1 ## ── Conflicts ────────────────────────────────────────── tidyverse_conflicts() ── ## ✖ tidyr::expand() masks Matrix::expand() ## ✖ dplyr::filter() masks stats::filter() ## ✖ dplyr::lag() masks stats::lag() ## ✖ purrr::map() masks mixOmics::map() ## ✖ tidyr::pack() masks Matrix::pack() ## ✖ dplyr::select() masks MASS::select() ## ✖ tidyr::unpack() masks Matrix::unpack() ## ℹ Use the conflicted package () to force all conflicts to become errors ``` r library(RVAideMemoire) ``` ## *** Package RVAideMemoire v 0.9-83-7 *** ## ## Attaching package: 'RVAideMemoire' ## ## The following object is masked from 'package:broom': ## ## bootstrap ## ## The following object is masked from 'package:lme4': ## ## dummy ``` r library(Hmisc) ``` ## ## Attaching package: 'Hmisc' ## ## The following objects are masked from 'package:dplyr': ## ## src, summarize ## ## The following objects are masked from 'package:base': ## ## format.pval, units ``` r library(corrplot) ``` ## corrplot 0.92 loaded # Read in gene count matrix Gene count matrix for all genes ``` r # raw gene counts data (will filter and variance stabilize) Peve_genes <- read_csv("E-Peve/output/02.20-E-Peve-RNAseq-alignment-HiSat2/peve-gene_count_matrix.csv") ``` ## Rows: 40389 Columns: 39 ## ── Column specification ──────────────────────────────────────────────────────── ## Delimiter: "," ## chr (1): gene_id ## dbl (38): POR-216-TP1, POR-216-TP2, POR-216-TP3, POR-216-TP4, POR-236-TP1, P... ## ## ℹ Use `spec()` to retrieve the full column specification for this data. ## ℹ Specify the column types or set `show_col_types = FALSE` to quiet this message. ``` r Peve_genes <- as.data.frame(Peve_genes) #Remove the "-gene" prefix from the gene ID column Peve_genes<-Peve_genes%>% mutate(gene_id = str_remove(gene_id, "^gene-"))%>% as.data.frame() # format gene IDs as rownames (instead of a column) rownames(Peve_genes) <- Peve_genes$gene_id Peve_genes <- Peve_genes%>%dplyr::select(!gene_id) colnames(Peve_genes) <- str_replace(colnames(Peve_genes), "^([^\\-]+\\-[^\\-]+)\\-", "\\1_") # load and format metadata metadata <- read_csv("M-multi-species/data/rna_metadata.csv")%>%dplyr::select(AzentaSampleName, ColonyID, Timepoint) %>% filter(grepl("POR", ColonyID)) ``` ## New names: ## Rows: 117 Columns: 19 ## ── Column specification ## ──────────────────────────────────────────────────────── Delimiter: "," chr ## (13): SampleName, WellNumber, AzentaSampleName, ColonyID, Timepoint, Sam... dbl ## (5): SampleNumber, Plate, TotalAmount-ng, Volume-uL, Conc-ng.uL lgl (1): ## MethodUsedForSpectrophotometry ## ℹ Use `spec()` to retrieve the full column specification for this data. ℹ ## Specify the column types or set `show_col_types = FALSE` to quiet this message. ## • `` -> `...19` ``` r metadata$Sample <- paste(metadata$ColonyID, metadata$Timepoint, sep = "_") colonies <- unique(metadata$ColonyID) # Load physiological data phys<-read_csv("https://github.com/urol-e5/timeseries/raw/refs/heads/master/time_series_analysis/Output/master_timeseries.csv")%>%filter(colony_id_corr %in% colonies)%>% dplyr::select(colony_id_corr, species, timepoint, site, Host_AFDW.mg.cm2, Sym_AFDW.mg.cm2, Am, AQY, Rd, Ik, Ic, calc.umol.cm2.hr, cells.mgAFDW, prot_mg.mgafdw, Ratio_AFDW.mg.cm2, Total_Chl, Total_Chl_cell, cre.umol.mgafdw) ``` ## Rows: 448 Columns: 46 ## ── Column specification ──────────────────────────────────────────────────────── ## Delimiter: "," ## chr (10): colony_id, colony_id_corr, species, timepoint, month, site, nutrie... ## dbl (36): cre.umol.mgprot, Host_AFDW.mg.cm2, Sym_AFDW.mg.cm2, Host_DW.mg.cm2... ## ## ℹ Use `spec()` to retrieve the full column specification for this data. ## ℹ Specify the column types or set `show_col_types = FALSE` to quiet this message. ``` r # format timepoint phys$timepoint <- gsub("timepoint", "TP", phys$timepoint) #add column with full sample info phys <- merge(phys, metadata, by.x = c("colony_id_corr", "timepoint"), by.y = c("ColonyID", "Timepoint")) %>% dplyr::select(-AzentaSampleName) #add site information into metadata metadata$Site<-phys$site[match(metadata$ColonyID, phys$colony_id_corr)] # Rename gene column names to include full sample info (as in miRNA table) #colnames(Apul_genes) <- metadata$Sample[match(colnames(Apul_genes), metadata$AzentaSampleName)] ``` # Gene set 1: Glycolysis Load in gene set generated by Peve-energy-go script ``` r glycolysis_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0006096_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000079 = col_character(), ## `sp|O97562|UCP2_PIG` = col_character(), ## `35.106` = col_double(), ## `282` = col_double(), ## `167` = col_double(), ## `5` = col_double(), ## `33` = col_double(), ## `301` = col_double(), ## `17` = col_double(), ## `295` = col_double(), ## `6.46e-49` = col_double(), ## `163` = col_double() ## ) Subset gene count matrix for this gene set. ``` r glycolysis_genes<-Peve_genes%>% filter(rownames(.) %in% glycolysis_go) ``` Calculate the sum of the total gene set for each sample. ``` r glycolysis_genes<-as.data.frame(t(glycolysis_genes)) glycolysis_genes$Sample<-rownames(glycolysis_genes) glycolysis_genes<-glycolysis_genes %>% rowwise() %>% mutate(glycolysis_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data<-left_join(phys, glycolysis_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data%>% ggplot(aes(x=timepoint, y=glycolysis_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-7-1.png) Plot as a PCA. ``` r pca_data <- data %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 788.2 0.10598 1.3434 0.119 ## Residual 34 6648.8 0.89402 ## Total 37 7437.0 1.00000 View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-11-1.png) Are there genes different by time point? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "Glycolysis", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-12-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00033963 2.29727741 ## Peve_00038087 2.06300202 ## Peve_00024602 1.95676921 ## Peve_00016096 1.94733180 ## Peve_00020350 1.91657065 ## Peve_00029657 1.78154850 ## Peve_00033950 1.76687439 ## Peve_00004659 1.74481509 ## Peve_00026350 1.74398003 ## Peve_00003158 1.72667526 ## Peve_00001586 1.67965218 ## Peve_00015643 1.67670872 ## Peve_00025794 1.63174479 ## Peve_00028656 1.61599470 ## Peve_00037310 1.54796932 ## Peve_00016268 1.53106537 ## Peve_00029032 1.50622400 ## Peve_00026729 1.41932988 ## Peve_00016809 1.41729244 ## Peve_00012722 1.38216340 ## Peve_00044760 1.37998883 ## Peve_00043390 1.37411309 ## Peve_00026977 1.36428046 ## Peve_00040401 1.34116034 ## Peve_00027083 1.33672977 ## Peve_00021856 1.33596835 ## Peve_00031790 1.33507836 ## Peve_00013838 1.33305138 ## Peve_00042157 1.32407468 ## Peve_00003539 1.32005827 ## Peve_00019806 1.31190308 ## Peve_00027073 1.30574730 ## Peve_00020572 1.29416791 ## Peve_00015045 1.27937656 ## Peve_00029654 1.27772162 ## Peve_00016646 1.27059237 ## Peve_00021245 1.26795430 ## Peve_00022328 1.26591970 ## Peve_00014624 1.26370595 ## Peve_00033476 1.25124486 ## Peve_00015518 1.21635205 ## Peve_00022867 1.20615562 ## Peve_00044062 1.20519140 ## Peve_00002321 1.20417487 ## Peve_00026728 1.19914806 ## Peve_00044421 1.19182653 ## Peve_00012020 1.18106763 ## Peve_00040733 1.17495078 ## Peve_00031785 1.16330283 ## Peve_00001411 1.15433075 ## Peve_00003570 1.14891602 ## Peve_00033089 1.14663160 ## Peve_00007243 1.13731246 ## Peve_00009560 1.13320289 ## Peve_00042302 1.12571563 ## Peve_00014159 1.12157060 ## Peve_00000114 1.10041690 ## Peve_00003219 1.09284168 ## Peve_00022830 1.08040741 ## Peve_00009073 1.07979774 ## Peve_00019240 1.07510531 ## Peve_00016361 1.06835306 ## Peve_00005527 1.06410905 ## Peve_00031326 1.06209340 ## Peve_00001435 1.05929942 ## Peve_00007365 1.05756829 ## Peve_00004657 1.05496814 ## Peve_00019361 1.05477124 ## Peve_00044666 1.04717944 ## Peve_00040519 1.03765074 ## Peve_00017602 1.03509939 ## Peve_00018570 1.02670854 ## Peve_00017071 1.02170343 ## Peve_00012273 0.99329999 ## Peve_00004660 0.98422554 ## Peve_00026347 0.97632261 ## Peve_00005352 0.97483136 ## Peve_00007726 0.97198678 ## Peve_00010022 0.97177005 ## Peve_00031003 0.96025805 ## Peve_00023694 0.95906842 ## Peve_00029178 0.95160027 ## Peve_00020457 0.94676473 ## Peve_00023464 0.94499178 ## Peve_00028171 0.93992452 ## Peve_00027572 0.93594819 ## Peve_00012587 0.93093792 ## Peve_00036445 0.92949213 ## Peve_00018899 0.92835309 ## Peve_00000418 0.92654947 ## Peve_00032593 0.91878755 ## Peve_00045066 0.91330935 ## Peve_00039119 0.90365776 ## Peve_00007764 0.89155469 ## Peve_00034319 0.88963361 ## Peve_00018336 0.88547219 ## Peve_00033645 0.88287420 ## Peve_00021861 0.87973173 ## Peve_00028793 0.85779089 ## Peve_00041828 0.85170788 ## Peve_00038394 0.84935335 ## Peve_00042168 0.84913127 ## Peve_00008951 0.84790214 ## Peve_00014328 0.84725291 ## Peve_00002969 0.84055710 ## Peve_00040732 0.83754441 ## Peve_00041779 0.83707702 ## Peve_00033826 0.83675524 ## Peve_00022156 0.83461986 ## Peve_00025321 0.83357023 ## Peve_00040503 0.83275613 ## Peve_00007364 0.82699702 ## Peve_00044998 0.82384087 ## Peve_00031599 0.81776057 ## Peve_00041056 0.81674940 ## Peve_00025384 0.81638612 ## Peve_00027369 0.81229141 ## Peve_00037725 0.81066756 ## Peve_00017515 0.79335512 ## Peve_00003792 0.79299457 ## Peve_00042763 0.78856815 ## Peve_00044664 0.78482127 ## Peve_00009969 0.77985286 ## Peve_00039118 0.77647505 ## Peve_00000391 0.77176008 ## Peve_00041037 0.77117294 ## Peve_00036957 0.76925045 ## Peve_00019553 0.76470224 ## Peve_00029726 0.75386195 ## Peve_00033436 0.75302077 ## Peve_00026674 0.74645725 ## Peve_00042762 0.74539695 ## Peve_00020199 0.74405144 ## Peve_00004630 0.74394847 ## Peve_00039201 0.73754216 ## Peve_00014688 0.72230857 ## Peve_00036372 0.72214391 ## Peve_00006937 0.71818044 ## Peve_00038665 0.71145995 ## Peve_00017049 0.70533003 ## Peve_00008952 0.69582006 ## Peve_00000209 0.69459855 ## Peve_00009808 0.69321089 ## Peve_00007821 0.68534053 ## Peve_00027717 0.68181204 ## Peve_00032931 0.66620306 ## Peve_00022742 0.65317425 ## Peve_00016095 0.65236011 ## Peve_00016743 0.64479308 ## Peve_00035951 0.64290673 ## Peve_00032222 0.64152352 ## Peve_00008881 0.63869412 ## Peve_00016589 0.63720891 ## Peve_00004978 0.62801938 ## Peve_00031001 0.62045578 ## Peve_00037749 0.61349046 ## Peve_00028934 0.61329257 ## Peve_00006921 0.61129067 ## Peve_00039585 0.60983438 ## Peve_00024151 0.60780500 ## Peve_00025007 0.60409894 ## Peve_00045154 0.59550888 ## Peve_00026737 0.59476138 ## Peve_00018002 0.58787463 ## Peve_00002770 0.58640462 ## Peve_00032223 0.58467663 ## Peve_00018571 0.56806361 ## Peve_00025006 0.55407112 ## Peve_00003538 0.54230798 ## Peve_00023980 0.54077972 ## Peve_00039845 0.52661165 ## Peve_00006977 0.52594123 ## Peve_00033827 0.52404903 ## Peve_00005351 0.51876790 ## Peve_00018895 0.50934669 ## Peve_00004634 0.50396802 ## Peve_00025848 0.50243441 ## Peve_00031783 0.49770142 ## Peve_00009866 0.49120238 ## Peve_00020608 0.48899272 ## Peve_00034834 0.48606815 ## Peve_00039958 0.48056216 ## Peve_00018723 0.46643487 ## Peve_00028378 0.45124312 ## Peve_00004071 0.44907168 ## Peve_00012559 0.44808404 ## Peve_00013429 0.42941765 ## Peve_00035358 0.41951844 ## Peve_00025417 0.41662198 ## Peve_00011363 0.40830199 ## Peve_00031596 0.38134584 ## Peve_00006976 0.37932686 ## Peve_00004025 0.34541396 ## Peve_00010476 0.34119690 ## Peve_00024621 0.30971680 ## Peve_00025500 0.28411450 ## Peve_00045007 0.23024002 ## Peve_00040465 0.22654523 ## Peve_00031151 0.19706388 ## Peve_00005980 0.17127345 ## Peve_00001733 0.03048582 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Glycolysis") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-13-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00017071 1.021703 ## 2 Peve_00018570 1.026709 ## 3 Peve_00017602 1.035099 ## 4 Peve_00040519 1.037651 ## 5 Peve_00044666 1.047179 ## 6 Peve_00019361 1.054771 ## 7 Peve_00004657 1.054968 ## 8 Peve_00007365 1.057568 ## 9 Peve_00001435 1.059299 ## 10 Peve_00031326 1.062093 ## 11 Peve_00005527 1.064109 ## 12 Peve_00016361 1.068353 ## 13 Peve_00019240 1.075105 ## 14 Peve_00009073 1.079798 ## 15 Peve_00022830 1.080407 ## 16 Peve_00003219 1.092842 ## 17 Peve_00000114 1.100417 ## 18 Peve_00014159 1.121571 ## 19 Peve_00042302 1.125716 ## 20 Peve_00009560 1.133203 ## 21 Peve_00007243 1.137312 ## 22 Peve_00033089 1.146632 ## 23 Peve_00003570 1.148916 ## 24 Peve_00001411 1.154331 ## 25 Peve_00031785 1.163303 ## 26 Peve_00040733 1.174951 ## 27 Peve_00012020 1.181068 ## 28 Peve_00044421 1.191827 ## 29 Peve_00026728 1.199148 ## 30 Peve_00002321 1.204175 ## 31 Peve_00044062 1.205191 ## 32 Peve_00022867 1.206156 ## 33 Peve_00015518 1.216352 ## 34 Peve_00033476 1.251245 ## 35 Peve_00014624 1.263706 ## 36 Peve_00022328 1.265920 ## 37 Peve_00021245 1.267954 ## 38 Peve_00016646 1.270592 ## 39 Peve_00029654 1.277722 ## 40 Peve_00015045 1.279377 ## 41 Peve_00020572 1.294168 ## 42 Peve_00027073 1.305747 ## 43 Peve_00019806 1.311903 ## 44 Peve_00003539 1.320058 ## 45 Peve_00042157 1.324075 ## 46 Peve_00013838 1.333051 ## 47 Peve_00031790 1.335078 ## 48 Peve_00021856 1.335968 ## 49 Peve_00027083 1.336730 ## 50 Peve_00040401 1.341160 ## 51 Peve_00026977 1.364280 ## 52 Peve_00043390 1.374113 ## 53 Peve_00044760 1.379989 ## 54 Peve_00012722 1.382163 ## 55 Peve_00016809 1.417292 ## 56 Peve_00026729 1.419330 ## 57 Peve_00029032 1.506224 ## 58 Peve_00016268 1.531065 ## 59 Peve_00037310 1.547969 ## 60 Peve_00028656 1.615995 ## 61 Peve_00025794 1.631745 ## 62 Peve_00015643 1.676709 ## 63 Peve_00001586 1.679652 ## 64 Peve_00003158 1.726675 ## 65 Peve_00026350 1.743980 ## 66 Peve_00004659 1.744815 ## 67 Peve_00033950 1.766874 ## 68 Peve_00029657 1.781548 ## 69 Peve_00020350 1.916571 ## 70 Peve_00016096 1.947332 ## 71 Peve_00024602 1.956769 ## 72 Peve_00038087 2.063002 ## 73 Peve_00033963 2.297277 Plot important genes ``` r plot<-data%>% ggplot(aes(x=timepoint, y=Peve_00033963, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-14-1.png) Plot second most important. ``` r plot<-data%>% ggplot(aes(x=timepoint, y=Peve_00038087, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-15-1.png) Plot third most important. ``` r plot<-data%>% ggplot(aes(x=timepoint, y=Peve_00024602, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-16-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 356.58 0.13202 1.7238 0.034 * ## Residual 34 2344.42 0.86798 ## Total 37 2701.00 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Glycolysis")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-20-1.png) Pull out PC1 score for each sample for GO term. ``` r scores <- scaled.pca$x scores<-as.data.frame(scores) scores<-scores%>%dplyr::select(PC1) scores$sample<-pca_data_vips$colony_id_corr scores$timepoint<-pca_data_vips$timepoint scores<-scores%>% rename(glycolysis=PC1) head(scores) ``` ## glycolysis sample timepoint ## 1 -3.423127 POR-216 TP1 ## 2 -13.074547 POR-216 TP2 ## 3 1.186372 POR-216 TP3 ## 4 4.455591 POR-216 TP4 ## 5 -9.380917 POR-236 TP1 ## 6 1.519258 POR-236 TP2 # Gene set 2: Gluconeogenesis Load in gene set generated by Apul-energy-go script ``` r gluconeo_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0006094_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000079 = col_character(), ## `sp|Q9UBX3|DIC_HUMAN` = col_character(), ## `61.348` = col_double(), ## `282` = col_double(), ## `106` = col_double(), ## `3` = col_double(), ## `27` = col_double(), ## `307` = col_double(), ## `5` = col_double(), ## `284` = col_double(), ## `9.30e-128` = col_double(), ## `363` = col_double() ## ) Subset gene count matrix for this gene set. ``` r gluconeo_genes<-Peve_genes%>% filter(rownames(.) %in% gluconeo_go) ``` Calculate the sum of the total gene set for each sample. ``` r gluconeo_genes<-as.data.frame(t(gluconeo_genes)) gluconeo_genes$Sample<-rownames(gluconeo_genes) gluconeo_genes<-gluconeo_genes %>% rowwise() %>% mutate(gluconeo_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data2<-left_join(phys, gluconeo_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data2%>% ggplot(aes(x=timepoint, y=gluconeo_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-26-1.png) Plot as a PCA. ``` r pca_data <- data2 %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 902.2 0.1318 1.7205 0.04 * ## Residual 34 5942.8 0.8682 ## Total 37 6845.0 1.0000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-30-1.png) Which genes are driving this? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "Gluconeogenesis", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-31-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00003523 2.2020393 ## Peve_00021850 1.8184345 ## Peve_00017775 1.8164215 ## Peve_00037415 1.7509739 ## Peve_00040632 1.5750100 ## Peve_00031000 1.5503151 ## Peve_00027763 1.4954287 ## Peve_00033369 1.4934716 ## Peve_00012132 1.4878704 ## Peve_00030742 1.4559505 ## Peve_00002053 1.4409764 ## Peve_00005205 1.4011433 ## Peve_00004922 1.3948744 ## Peve_00029370 1.3901128 ## Peve_00003158 1.3892511 ## Peve_00006508 1.3841796 ## Peve_00028672 1.3707337 ## Peve_00040399 1.3438049 ## Peve_00022915 1.3392245 ## Peve_00032681 1.3247457 ## Peve_00003003 1.3144545 ## Peve_00035350 1.3139105 ## Peve_00002770 1.3054433 ## Peve_00015643 1.3018288 ## Peve_00017576 1.2881001 ## Peve_00022082 1.2647923 ## Peve_00038388 1.2572085 ## Peve_00032890 1.2550320 ## Peve_00017459 1.2538755 ## Peve_00028481 1.2535505 ## Peve_00043694 1.2447196 ## Peve_00035492 1.2360014 ## Peve_00011193 1.2341325 ## Peve_00027796 1.2340782 ## Peve_00003879 1.2276343 ## Peve_00016361 1.2252368 ## Peve_00008487 1.2218106 ## Peve_00040520 1.2042993 ## Peve_00016285 1.2001786 ## Peve_00021477 1.1882022 ## Peve_00041570 1.1869318 ## Peve_00036485 1.1867027 ## Peve_00004469 1.1814521 ## Peve_00021031 1.1805823 ## Peve_00028811 1.1760254 ## Peve_00038364 1.1690568 ## Peve_00034684 1.1662316 ## Peve_00029852 1.1546701 ## Peve_00017777 1.1512344 ## Peve_00037340 1.1501076 ## Peve_00035574 1.1433581 ## Peve_00040413 1.1261282 ## Peve_00019553 1.1220267 ## Peve_00014226 1.1143988 ## Peve_00003533 1.1124688 ## Peve_00043390 1.1119368 ## Peve_00012220 1.1109683 ## Peve_00028171 1.0786618 ## Peve_00018002 1.0780391 ## Peve_00035371 1.0746720 ## Peve_00036382 1.0726754 ## Peve_00038365 1.0605132 ## Peve_00023586 1.0577297 ## Peve_00040519 1.0572593 ## Peve_00042417 1.0539271 ## Peve_00017613 1.0409495 ## Peve_00008862 1.0383614 ## Peve_00006258 1.0375157 ## Peve_00039255 1.0350023 ## Peve_00018649 1.0237334 ## Peve_00042157 1.0226243 ## Peve_00012228 1.0202052 ## Peve_00007726 1.0143311 ## Peve_00008236 1.0128945 ## Peve_00017602 1.0123965 ## Peve_00029395 1.0041935 ## Peve_00012587 0.9918678 ## Peve_00003528 0.9908753 ## Peve_00041242 0.9811291 ## Peve_00010044 0.9723435 ## Peve_00019240 0.9676238 ## Peve_00010033 0.9651402 ## Peve_00003361 0.9620201 ## Peve_00044663 0.9619335 ## Peve_00009730 0.9613775 ## Peve_00034371 0.9575052 ## Peve_00008103 0.9555568 ## Peve_00026674 0.9455668 ## Peve_00012433 0.9439027 ## Peve_00012273 0.9417393 ## Peve_00033645 0.9287986 ## Peve_00003439 0.9246893 ## Peve_00032896 0.9183120 ## Peve_00023588 0.9105777 ## Peve_00032861 0.9097257 ## Peve_00031452 0.9076593 ## Peve_00037419 0.9046973 ## Peve_00037070 0.8922609 ## Peve_00006000 0.8887866 ## Peve_00034681 0.8878743 ## Peve_00009726 0.8855778 ## Peve_00008231 0.8833475 ## Peve_00037278 0.8715987 ## Peve_00037817 0.8649846 ## Peve_00014710 0.8582113 ## Peve_00002148 0.8566428 ## Peve_00002050 0.8530950 ## Peve_00019809 0.8513851 ## Peve_00041056 0.8505226 ## Peve_00015579 0.8498295 ## Peve_00040552 0.8489469 ## Peve_00034834 0.8426395 ## Peve_00014328 0.8403748 ## Peve_00039187 0.8321441 ## Peve_00009969 0.8311370 ## Peve_00023587 0.8276228 ## Peve_00004462 0.8219470 ## Peve_00030392 0.8213284 ## Peve_00031630 0.8211397 ## Peve_00016245 0.8205044 ## Peve_00005211 0.8161727 ## Peve_00037180 0.8147067 ## Peve_00004978 0.8107594 ## Peve_00006230 0.8104648 ## Peve_00000418 0.8096330 ## Peve_00027369 0.8083231 ## Peve_00010690 0.8024150 ## Peve_00040946 0.8006128 ## Peve_00039004 0.7910773 ## Peve_00033089 0.7865846 ## Peve_00039032 0.7830092 ## Peve_00007270 0.7810574 ## Peve_00043808 0.7806844 ## Peve_00003529 0.7758200 ## Peve_00004308 0.7529076 ## Peve_00028667 0.7496030 ## Peve_00043038 0.7472513 ## Peve_00025819 0.7455804 ## Peve_00002890 0.7446924 ## Peve_00010172 0.7442618 ## Peve_00022633 0.7385173 ## Peve_00017238 0.7264070 ## Peve_00034411 0.7246432 ## Peve_00020490 0.7173064 ## Peve_00004071 0.7143102 ## Peve_00004468 0.7123780 ## Peve_00027370 0.7080572 ## Peve_00006507 0.7058833 ## Peve_00017262 0.6953082 ## Peve_00037724 0.6923009 ## Peve_00005350 0.6900063 ## Peve_00034442 0.6787867 ## Peve_00037181 0.6687205 ## Peve_00031001 0.6641380 ## Peve_00027710 0.6608170 ## Peve_00013532 0.6585120 ## Peve_00012434 0.6573702 ## Peve_00032354 0.6443032 ## Peve_00026725 0.6436848 ## Peve_00013255 0.6319186 ## Peve_00043673 0.6291934 ## Peve_00025258 0.6204576 ## Peve_00003570 0.6069976 ## Peve_00040401 0.6030500 ## Peve_00022742 0.5948961 ## Peve_00019384 0.5729757 ## Peve_00045074 0.5647421 ## Peve_00023585 0.5504449 ## Peve_00002534 0.5468384 ## Peve_00019361 0.5385266 ## Peve_00023281 0.5080967 ## Peve_00027579 0.4944957 ## Peve_00021324 0.4939870 ## Peve_00009647 0.4821455 ## Peve_00020571 0.4641428 ## Peve_00028673 0.4305067 ## Peve_00038787 0.4281249 ## Peve_00018569 0.4093585 ## Peve_00024576 0.3962509 ## Peve_00023823 0.3819573 ## Peve_00037417 0.3702621 ## Peve_00024621 0.3625989 ## Peve_00045157 0.3597447 ## Peve_00020576 0.3493728 ## Peve_00035418 0.2601230 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Gluconeogenesis") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-32-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00029395 1.004194 ## 2 Peve_00017602 1.012397 ## 3 Peve_00008236 1.012895 ## 4 Peve_00007726 1.014331 ## 5 Peve_00012228 1.020205 ## 6 Peve_00042157 1.022624 ## 7 Peve_00018649 1.023733 ## 8 Peve_00039255 1.035002 ## 9 Peve_00006258 1.037516 ## 10 Peve_00008862 1.038361 ## 11 Peve_00017613 1.040950 ## 12 Peve_00042417 1.053927 ## 13 Peve_00040519 1.057259 ## 14 Peve_00023586 1.057730 ## 15 Peve_00038365 1.060513 ## 16 Peve_00036382 1.072675 ## 17 Peve_00035371 1.074672 ## 18 Peve_00018002 1.078039 ## 19 Peve_00028171 1.078662 ## 20 Peve_00012220 1.110968 ## 21 Peve_00043390 1.111937 ## 22 Peve_00003533 1.112469 ## 23 Peve_00014226 1.114399 ## 24 Peve_00019553 1.122027 ## 25 Peve_00040413 1.126128 ## 26 Peve_00035574 1.143358 ## 27 Peve_00037340 1.150108 ## 28 Peve_00017777 1.151234 ## 29 Peve_00029852 1.154670 ## 30 Peve_00034684 1.166232 ## 31 Peve_00038364 1.169057 ## 32 Peve_00028811 1.176025 ## 33 Peve_00021031 1.180582 ## 34 Peve_00004469 1.181452 ## 35 Peve_00036485 1.186703 ## 36 Peve_00041570 1.186932 ## 37 Peve_00021477 1.188202 ## 38 Peve_00016285 1.200179 ## 39 Peve_00040520 1.204299 ## 40 Peve_00008487 1.221811 ## 41 Peve_00016361 1.225237 ## 42 Peve_00003879 1.227634 ## 43 Peve_00027796 1.234078 ## 44 Peve_00011193 1.234133 ## 45 Peve_00035492 1.236001 ## 46 Peve_00043694 1.244720 ## 47 Peve_00028481 1.253551 ## 48 Peve_00017459 1.253876 ## 49 Peve_00032890 1.255032 ## 50 Peve_00038388 1.257209 ## 51 Peve_00022082 1.264792 ## 52 Peve_00017576 1.288100 ## 53 Peve_00015643 1.301829 ## 54 Peve_00002770 1.305443 ## 55 Peve_00035350 1.313910 ## 56 Peve_00003003 1.314455 ## 57 Peve_00032681 1.324746 ## 58 Peve_00022915 1.339224 ## 59 Peve_00040399 1.343805 ## 60 Peve_00028672 1.370734 ## 61 Peve_00006508 1.384180 ## 62 Peve_00003158 1.389251 ## 63 Peve_00029370 1.390113 ## 64 Peve_00004922 1.394874 ## 65 Peve_00005205 1.401143 ## 66 Peve_00002053 1.440976 ## 67 Peve_00030742 1.455951 ## 68 Peve_00012132 1.487870 ## 69 Peve_00033369 1.493472 ## 70 Peve_00027763 1.495429 ## 71 Peve_00031000 1.550315 ## 72 Peve_00040632 1.575010 ## 73 Peve_00037415 1.750974 ## 74 Peve_00017775 1.816422 ## 75 Peve_00021850 1.818434 ## 76 Peve_00003523 2.202039 Plot top genes. ``` r plot<-data2%>% ggplot(aes(x=timepoint, y=Peve_00003523, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-33-1.png) Plot second most important. ``` r plot<-data2%>% ggplot(aes(x=timepoint, y=Peve_00021850, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-34-1.png) Plot third most important. ``` r plot<-data2%>% ggplot(aes(x=timepoint, y=Peve_00017775, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-35-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 509.17 0.18107 2.5059 0.007 ** ## Residual 34 2302.83 0.81893 ## Total 37 2812.00 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Gluconeogenesis")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-39-1.png) Pull out PC1 score for each sample for GO term. ``` r scores1 <- scaled.pca$x scores1<-as.data.frame(scores1) scores1<-scores1%>%dplyr::select(PC1) scores1$sample<-pca_data_vips$colony_id_corr scores1$timepoint<-pca_data_vips$timepoint scores1<-scores1%>% rename(gluconeogenesis=PC1) scores<-left_join(scores, scores1) ``` ## Joining with `by = join_by(sample, timepoint)` ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis ## 1 -3.423127 POR-216 TP1 -6.749172 ## 2 -13.074547 POR-216 TP2 -10.860762 ## 3 1.186372 POR-216 TP3 1.457899 ## 4 4.455591 POR-216 TP4 7.899062 ## 5 -9.380917 POR-236 TP1 -11.102232 ## 6 1.519258 POR-236 TP2 3.529447 # Gene set 3: Lipolysis/lipid catabolism Load in gene set generated by Apul-energy-go script ``` r lipolysis_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0016042_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000150 = col_character(), ## `sp|O49561|G2OX8_ARATH` = col_character(), ## `30.000` = col_double(), ## `280` = col_double(), ## `168` = col_double(), ## `8` = col_double(), ## `3` = col_double(), ## `279` = col_double(), ## `42` = col_double(), ## `296` = col_double(), ## `8.27e-32` = col_double(), ## `119` = col_double() ## ) Subset gene count matrix for this gene set. ``` r lipolysis_genes<-Peve_genes%>% filter(rownames(.) %in% lipolysis_go) ``` Calculate the sum of the total gene set for each sample. ``` r lipolysis_genes<-as.data.frame(t(lipolysis_genes)) lipolysis_genes$Sample<-rownames(lipolysis_genes) lipolysis_genes<-lipolysis_genes %>% rowwise() %>% mutate(lipolysis_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data3<-left_join(phys, lipolysis_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=lipolysis_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-45-1.png) Plot as a PCA. ``` r pca_data <- data3 %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 4229 0.1038 1.3127 0.115 ## Residual 34 36508 0.8962 ## Total 37 40737 1.0000 View by timepoint ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=FALSE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-49-1.png) Which genes are driving this? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "Lipolysis", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-50-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00017775 2.1715182 ## Peve_00006804 2.1131345 ## Peve_00021850 2.1053841 ## Peve_00032336 2.0882591 ## Peve_00033018 2.0061051 ## Peve_00005936 1.9964799 ## Peve_00038430 1.9850825 ## Peve_00001961 1.9045622 ## Peve_00023487 1.8969505 ## Peve_00036049 1.8724582 ## Peve_00037585 1.8436706 ## Peve_00036939 1.8355605 ## Peve_00022762 1.8027845 ## Peve_00021880 1.7903000 ## Peve_00032856 1.7860496 ## Peve_00016032 1.7793200 ## Peve_00036046 1.7776571 ## Peve_00038749 1.7611318 ## Peve_00027198 1.7350277 ## Peve_00004624 1.7322801 ## Peve_00006753 1.7200365 ## Peve_00015834 1.6788684 ## Peve_00001194 1.6772963 ## Peve_00008664 1.6742312 ## Peve_00028037 1.6595831 ## Peve_00026350 1.6557498 ## Peve_00006892 1.6475433 ## Peve_00030037 1.6451702 ## Peve_00017203 1.6381495 ## Peve_00030742 1.6377827 ## Peve_00000626 1.6359788 ## Peve_00002968 1.6358869 ## Peve_00034757 1.6308602 ## Peve_00016220 1.6305254 ## Peve_00019683 1.6272281 ## Peve_00023375 1.6231227 ## Peve_00006912 1.6218438 ## Peve_00032844 1.6157929 ## Peve_00024810 1.6139177 ## Peve_00002772 1.6074553 ## Peve_00021294 1.6059982 ## Peve_00024065 1.5945168 ## Peve_00022915 1.5940988 ## Peve_00034022 1.5925554 ## Peve_00031211 1.5900577 ## Peve_00028038 1.5796486 ## Peve_00033530 1.5793834 ## Peve_00043558 1.5680273 ## Peve_00011444 1.5596564 ## Peve_00013614 1.5551544 ## Peve_00003252 1.5514351 ## Peve_00001522 1.5472004 ## Peve_00024386 1.5408633 ## Peve_00000528 1.5336760 ## Peve_00044494 1.5332297 ## Peve_00031844 1.5301144 ## Peve_00037635 1.5301040 ## Peve_00030007 1.5257571 ## Peve_00017897 1.5182101 ## Peve_00026126 1.5144292 ## Peve_00010859 1.5133882 ## Peve_00042079 1.5117567 ## Peve_00024967 1.5109261 ## Peve_00030297 1.5081580 ## Peve_00012723 1.5079175 ## Peve_00044383 1.5061728 ## Peve_00009113 1.5048237 ## Peve_00018372 1.5017945 ## Peve_00042832 1.4957882 ## Peve_00018677 1.4949186 ## Peve_00031366 1.4884081 ## Peve_00044100 1.4868638 ## Peve_00012936 1.4829600 ## Peve_00038751 1.4755650 ## Peve_00042919 1.4737674 ## Peve_00043434 1.4712192 ## Peve_00008132 1.4666190 ## Peve_00001052 1.4658288 ## Peve_00036567 1.4651181 ## Peve_00043616 1.4647010 ## Peve_00003535 1.4631164 ## Peve_00004242 1.4612056 ## Peve_00016809 1.4606602 ## Peve_00039167 1.4585869 ## Peve_00035492 1.4585364 ## Peve_00039635 1.4550056 ## Peve_00000818 1.4514970 ## Peve_00010277 1.4472931 ## Peve_00034700 1.4468666 ## Peve_00017766 1.4466725 ## Peve_00034484 1.4406832 ## Peve_00038364 1.4399183 ## Peve_00009162 1.4393518 ## Peve_00038504 1.4383846 ## Peve_00042077 1.4358619 ## Peve_00032855 1.4341865 ## Peve_00031923 1.4295511 ## Peve_00036458 1.4275185 ## Peve_00011089 1.4264479 ## Peve_00002053 1.4255919 ## Peve_00012877 1.4179630 ## Peve_00041570 1.4162248 ## Peve_00007801 1.4149175 ## Peve_00011095 1.4073109 ## Peve_00001490 1.4025207 ## Peve_00021477 1.3981878 ## Peve_00018424 1.3936931 ## Peve_00000296 1.3931959 ## Peve_00001262 1.3902848 ## Peve_00026382 1.3885257 ## Peve_00004344 1.3808728 ## Peve_00004715 1.3799913 ## Peve_00038432 1.3798203 ## Peve_00016285 1.3794322 ## Peve_00034013 1.3771466 ## Peve_00037573 1.3770772 ## Peve_00008766 1.3761817 ## Peve_00005087 1.3749587 ## Peve_00032890 1.3734711 ## Peve_00019247 1.3726519 ## Peve_00033821 1.3664552 ## Peve_00025988 1.3649606 ## Peve_00040520 1.3641877 ## Peve_00010077 1.3606559 ## Peve_00034790 1.3603944 ## Peve_00008487 1.3551563 ## Peve_00011191 1.3549379 ## Peve_00026130 1.3546936 ## Peve_00012132 1.3536289 ## Peve_00017462 1.3532184 ## Peve_00024466 1.3505490 ## Peve_00033476 1.3502198 ## Peve_00027083 1.3488917 ## Peve_00024408 1.3480728 ## Peve_00013257 1.3472579 ## Peve_00039654 1.3455647 ## Peve_00033950 1.3450785 ## Peve_00033529 1.3449896 ## Peve_00014607 1.3449750 ## Peve_00006894 1.3444848 ## Peve_00036485 1.3442062 ## Peve_00033189 1.3404262 ## Peve_00011193 1.3394232 ## Peve_00019287 1.3387367 ## Peve_00019000 1.3380268 ## Peve_00026425 1.3370807 ## Peve_00026107 1.3369714 ## Peve_00033216 1.3305189 ## Peve_00041684 1.3282608 ## Peve_00024010 1.3268367 ## Peve_00031436 1.3260272 ## Peve_00004544 1.3253627 ## Peve_00028710 1.3234428 ## Peve_00030718 1.3206044 ## Peve_00038767 1.3189351 ## Peve_00021243 1.3188751 ## Peve_00016733 1.3166593 ## Peve_00021331 1.3134436 ## Peve_00016361 1.3131656 ## Peve_00005411 1.3129805 ## Peve_00040557 1.3102716 ## Peve_00021995 1.3075245 ## Peve_00004364 1.3051420 ## Peve_00018935 1.3009518 ## Peve_00037340 1.3007909 ## Peve_00042234 1.3005762 ## Peve_00028674 1.3003824 ## Peve_00020371 1.2991478 ## Peve_00019983 1.2972127 ## Peve_00003539 1.2968586 ## Peve_00023784 1.2962639 ## Peve_00037697 1.2910120 ## Peve_00042048 1.2879806 ## Peve_00043615 1.2868459 ## Peve_00044062 1.2849050 ## Peve_00009666 1.2836561 ## Peve_00031448 1.2834036 ## Peve_00033364 1.2831202 ## Peve_00010060 1.2830537 ## Peve_00004363 1.2813531 ## Peve_00042762 1.2782531 ## Peve_00023967 1.2772314 ## Peve_00014495 1.2765538 ## Peve_00043371 1.2738372 ## Peve_00040399 1.2721858 ## Peve_00044933 1.2706299 ## Peve_00013550 1.2691404 ## Peve_00016735 1.2687613 ## Peve_00036495 1.2663864 ## Peve_00043373 1.2643623 ## Peve_00010640 1.2613062 ## Peve_00029040 1.2596740 ## Peve_00044947 1.2589805 ## Peve_00018969 1.2584899 ## Peve_00003359 1.2572368 ## Peve_00029735 1.2526941 ## Peve_00042929 1.2523413 ## Peve_00033589 1.2488353 ## Peve_00033815 1.2481925 ## Peve_00042769 1.2441951 ## Peve_00016660 1.2429377 ## Peve_00044340 1.2428145 ## Peve_00033895 1.2418862 ## Peve_00026632 1.2413670 ## Peve_00025183 1.2410011 ## Peve_00014918 1.2407448 ## Peve_00032327 1.2384604 ## Peve_00026750 1.2363666 ## Peve_00015398 1.2352391 ## Peve_00007692 1.2337399 ## Peve_00014195 1.2330088 ## Peve_00014814 1.2329638 ## Peve_00008027 1.2319038 ## Peve_00021295 1.2316535 ## Peve_00041745 1.2310318 ## Peve_00026798 1.2294802 ## Peve_00001054 1.2274626 ## Peve_00043472 1.2272857 ## Peve_00001876 1.2251898 ## Peve_00001870 1.2250754 ## Peve_00036060 1.2229235 ## Peve_00008880 1.2206952 ## Peve_00004770 1.2206384 ## Peve_00016661 1.2190846 ## Peve_00043495 1.2155417 ## Peve_00017667 1.2150710 ## Peve_00035085 1.2131337 ## Peve_00028493 1.2115920 ## Peve_00005767 1.2111793 ## Peve_00011538 1.2107008 ## Peve_00021920 1.2090377 ## Peve_00039877 1.2035404 ## Peve_00018289 1.2016846 ## Peve_00017935 1.2014686 ## Peve_00022114 1.2005720 ## Peve_00044378 1.1999607 ## Peve_00027053 1.1991728 ## Peve_00022089 1.1981515 ## Peve_00004095 1.1967502 ## Peve_00008519 1.1940054 ## Peve_00001927 1.1936712 ## Peve_00024064 1.1923551 ## Peve_00016635 1.1915624 ## Peve_00021214 1.1904875 ## Peve_00008862 1.1904014 ## Peve_00041156 1.1903637 ## Peve_00000698 1.1900335 ## Peve_00014226 1.1894502 ## Peve_00038365 1.1877274 ## Peve_00003553 1.1876362 ## Peve_00018719 1.1864388 ## Peve_00025803 1.1838810 ## Peve_00021915 1.1826271 ## Peve_00031667 1.1820486 ## Peve_00000209 1.1818480 ## Peve_00006506 1.1802554 ## Peve_00025755 1.1802179 ## Peve_00037091 1.1770812 ## Peve_00007232 1.1755262 ## Peve_00034406 1.1745775 ## Peve_00033366 1.1719602 ## Peve_00006258 1.1710598 ## Peve_00018886 1.1699717 ## Peve_00031027 1.1692749 ## Peve_00026567 1.1672715 ## Peve_00042344 1.1637568 ## Peve_00005527 1.1635931 ## Peve_00025825 1.1624068 ## Peve_00006966 1.1597480 ## Peve_00017927 1.1579528 ## Peve_00004269 1.1571595 ## Peve_00023296 1.1569908 ## Peve_00028896 1.1567820 ## Peve_00033537 1.1563302 ## Peve_00034981 1.1552743 ## Peve_00004950 1.1552572 ## Peve_00026220 1.1538640 ## Peve_00022385 1.1537003 ## Peve_00042727 1.1533678 ## Peve_00018649 1.1529192 ## Peve_00014586 1.1518927 ## Peve_00044421 1.1508665 ## Peve_00023545 1.1461537 ## Peve_00021274 1.1456022 ## Peve_00006750 1.1448363 ## Peve_00017195 1.1447012 ## Peve_00020731 1.1441920 ## Peve_00015295 1.1424822 ## Peve_00037669 1.1418039 ## Peve_00020611 1.1386620 ## Peve_00021919 1.1382911 ## Peve_00042601 1.1381396 ## Peve_00021060 1.1357161 ## Peve_00024021 1.1346537 ## Peve_00044663 1.1327436 ## Peve_00014491 1.1323710 ## Peve_00013091 1.1316751 ## Peve_00017493 1.1316303 ## Peve_00009337 1.1296986 ## Peve_00014158 1.1292192 ## Peve_00043842 1.1288801 ## Peve_00004882 1.1281287 ## Peve_00024673 1.1269800 ## Peve_00041125 1.1262723 ## Peve_00041789 1.1259535 ## Peve_00019671 1.1250233 ## Peve_00004187 1.1246842 ## Peve_00021554 1.1197904 ## Peve_00024909 1.1188262 ## Peve_00016851 1.1165121 ## Peve_00028350 1.1148688 ## Peve_00021042 1.1121838 ## Peve_00007389 1.1114234 ## Peve_00037694 1.1110541 ## Peve_00001653 1.1109184 ## Peve_00024531 1.1106862 ## Peve_00023351 1.1104333 ## Peve_00023047 1.1100492 ## Peve_00011919 1.1095022 ## Peve_00020563 1.1087101 ## Peve_00030224 1.1067094 ## Peve_00037923 1.1066076 ## Peve_00039590 1.1056535 ## Peve_00013503 1.1045663 ## Peve_00016985 1.1042503 ## Peve_00010033 1.1015433 ## Peve_00019163 1.1014311 ## Peve_00021861 1.1013783 ## Peve_00002423 1.1008296 ## Peve_00037711 1.1003606 ## Peve_00016374 1.1003145 ## Peve_00031935 1.0994824 ## Peve_00020828 1.0988327 ## Peve_00022617 1.0972062 ## Peve_00042679 1.0945446 ## Peve_00031444 1.0938443 ## Peve_00034146 1.0934835 ## Peve_00014486 1.0934321 ## Peve_00045006 1.0916991 ## Peve_00041161 1.0911777 ## Peve_00004102 1.0903437 ## Peve_00015583 1.0902468 ## Peve_00037817 1.0890935 ## Peve_00021914 1.0870467 ## Peve_00023037 1.0861057 ## Peve_00015538 1.0854307 ## Peve_00000751 1.0853514 ## Peve_00044176 1.0848421 ## Peve_00028192 1.0836265 ## Peve_00037587 1.0830585 ## Peve_00039587 1.0820325 ## Peve_00029395 1.0805388 ## Peve_00010247 1.0802176 ## Peve_00041692 1.0786673 ## Peve_00013481 1.0780027 ## Peve_00044734 1.0776677 ## Peve_00032154 1.0771777 ## Peve_00025334 1.0771611 ## Peve_00018546 1.0771271 ## Peve_00015302 1.0767050 ## Peve_00038540 1.0758096 ## Peve_00033431 1.0757660 ## Peve_00004356 1.0754047 ## Peve_00001337 1.0740620 ## Peve_00031914 1.0734211 ## Peve_00017419 1.0730942 ## Peve_00000781 1.0727335 ## Peve_00001210 1.0727078 ## Peve_00016637 1.0704678 ## Peve_00000594 1.0703573 ## Peve_00024966 1.0687190 ## Peve_00040532 1.0685937 ## Peve_00021008 1.0683889 ## Peve_00033177 1.0678023 ## Peve_00035307 1.0674690 ## Peve_00010301 1.0667351 ## Peve_00023464 1.0663524 ## Peve_00002785 1.0661485 ## Peve_00021461 1.0648400 ## Peve_00035494 1.0625959 ## Peve_00031025 1.0623937 ## Peve_00027940 1.0620367 ## Peve_00036629 1.0605472 ## Peve_00018570 1.0598374 ## Peve_00021061 1.0597265 ## Peve_00044990 1.0593942 ## Peve_00019002 1.0585038 ## Peve_00023995 1.0571750 ## Peve_00007470 1.0569561 ## Peve_00007945 1.0557508 ## Peve_00033917 1.0546608 ## Peve_00007102 1.0541442 ## Peve_00002214 1.0541155 ## Peve_00009227 1.0538101 ## Peve_00035298 1.0535144 ## Peve_00017515 1.0533156 ## Peve_00007764 1.0531115 ## Peve_00019553 1.0530926 ## Peve_00037696 1.0511928 ## Peve_00029547 1.0508084 ## Peve_00026033 1.0506266 ## Peve_00043996 1.0505215 ## Peve_00011448 1.0496124 ## Peve_00003439 1.0488160 ## Peve_00009460 1.0486926 ## Peve_00042049 1.0472808 ## Peve_00007980 1.0455671 ## Peve_00029230 1.0445766 ## Peve_00039903 1.0442338 ## Peve_00018376 1.0436273 ## Peve_00032593 1.0436199 ## Peve_00003847 1.0436080 ## Peve_00009817 1.0434373 ## Peve_00022577 1.0428254 ## Peve_00031155 1.0424575 ## Peve_00003361 1.0423509 ## Peve_00015391 1.0393117 ## Peve_00021006 1.0382170 ## Peve_00001454 1.0375691 ## Peve_00033897 1.0363875 ## Peve_00020616 1.0357425 ## Peve_00018122 1.0341539 ## Peve_00007379 1.0333953 ## Peve_00002050 1.0325162 ## Peve_00038750 1.0314201 ## Peve_00024913 1.0298149 ## Peve_00019422 1.0292496 ## Peve_00011540 1.0287693 ## Peve_00015410 1.0285157 ## Peve_00016042 1.0273683 ## Peve_00031228 1.0264030 ## Peve_00022578 1.0256244 ## Peve_00021814 1.0246984 ## Peve_00035454 1.0238534 ## Peve_00008974 1.0234032 ## Peve_00002402 1.0230808 ## Peve_00001258 1.0220140 ## Peve_00032369 1.0208801 ## Peve_00020522 1.0203916 ## Peve_00001193 1.0187577 ## Peve_00003892 1.0163401 ## Peve_00004130 1.0162377 ## Peve_00010097 1.0162079 ## Peve_00016927 1.0158118 ## Peve_00020213 1.0155131 ## Peve_00001359 1.0149125 ## Peve_00033394 1.0142857 ## Peve_00031933 1.0135915 ## Peve_00028040 1.0134305 ## Peve_00038918 1.0128766 ## Peve_00003219 1.0128080 ## Peve_00000568 1.0128018 ## Peve_00028041 1.0099909 ## Peve_00025143 1.0094349 ## Peve_00016663 1.0084599 ## Peve_00030402 1.0083665 ## Peve_00043372 1.0066197 ## Peve_00032155 1.0060132 ## Peve_00036957 1.0052138 ## Peve_00002096 1.0042372 ## Peve_00034142 1.0042072 ## Peve_00040836 1.0041726 ## Peve_00042194 1.0011509 ## Peve_00019684 1.0000279 ## Peve_00010690 0.9999239 ## Peve_00022086 0.9999140 ## Peve_00034052 0.9992649 ## Peve_00001257 0.9982095 ## Peve_00037684 0.9975676 ## Peve_00025146 0.9967630 ## Peve_00008001 0.9952099 ## Peve_00022156 0.9951311 ## Peve_00017091 0.9950306 ## Peve_00005057 0.9947353 ## Peve_00039119 0.9947256 ## Peve_00038251 0.9940152 ## Peve_00040548 0.9938029 ## Peve_00025335 0.9931672 ## Peve_00000384 0.9919561 ## Peve_00013037 0.9914977 ## Peve_00019809 0.9909748 ## Peve_00039798 0.9904912 ## Peve_00004174 0.9902329 ## Peve_00025329 0.9896336 ## Peve_00002062 0.9894099 ## Peve_00026589 0.9888431 ## Peve_00041779 0.9886867 ## Peve_00006215 0.9884424 ## Peve_00028039 0.9861382 ## Peve_00042535 0.9857596 ## Peve_00011116 0.9855311 ## Peve_00017895 0.9855177 ## Peve_00034498 0.9851165 ## Peve_00036854 0.9833932 ## Peve_00039448 0.9833394 ## Peve_00018607 0.9828683 ## Peve_00012447 0.9827914 ## 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Peve_00033630 0.4565334 ## Peve_00039508 0.4552585 ## Peve_00003462 0.4516243 ## Peve_00000665 0.4479595 ## Peve_00018293 0.4455126 ## Peve_00002061 0.4453173 ## Peve_00010023 0.4409238 ## Peve_00020387 0.4318905 ## Peve_00024769 0.4236709 ## Peve_00029076 0.4200129 ## Peve_00036204 0.4188142 ## Peve_00027320 0.4158357 ## Peve_00034763 0.4151257 ## Peve_00026127 0.4120222 ## Peve_00044754 0.4110923 ## Peve_00028576 0.4105512 ## Peve_00011736 0.4074146 ## Peve_00017476 0.4046295 ## Peve_00026134 0.4046263 ## Peve_00006937 0.4027190 ## Peve_00024959 0.4024169 ## Peve_00005766 0.3982670 ## Peve_00024938 0.3964631 ## Peve_00036496 0.3964088 ## Peve_00022188 0.3955883 ## Peve_00019535 0.3945966 ## Peve_00015932 0.3926217 ## Peve_00044998 0.3822135 ## Peve_00033771 0.3806148 ## Peve_00023597 0.3745231 ## Peve_00030611 0.3739863 ## Peve_00032584 0.3738677 ## Peve_00026860 0.3737849 ## Peve_00021899 0.3698507 ## Peve_00011860 0.3680279 ## Peve_00019680 0.3664292 ## Peve_00044927 0.3653286 ## Peve_00033571 0.3640543 ## Peve_00033569 0.3627763 ## Peve_00030355 0.3600534 ## Peve_00001372 0.3579221 ## Peve_00004351 0.3509434 ## Peve_00009193 0.3452439 ## Peve_00000715 0.3439003 ## Peve_00041467 0.3417742 ## Peve_00013040 0.3406688 ## Peve_00008952 0.3394576 ## Peve_00033229 0.3393406 ## Peve_00024576 0.3369974 ## Peve_00011839 0.3352550 ## Peve_00008082 0.3324158 ## Peve_00019393 0.3294819 ## Peve_00017200 0.3276871 ## Peve_00009152 0.3214890 ## Peve_00038063 0.3135867 ## Peve_00010869 0.3117388 ## Peve_00014913 0.3109959 ## Peve_00028036 0.3104012 ## Peve_00028634 0.3079193 ## Peve_00038665 0.2997091 ## Peve_00013897 0.2988947 ## Peve_00036463 0.2962318 ## Peve_00007419 0.2949663 ## Peve_00026273 0.2944395 ## Peve_00029677 0.2929872 ## Peve_00025147 0.2900512 ## Peve_00010095 0.2896616 ## Peve_00041704 0.2860001 ## Peve_00010098 0.2805614 ## Peve_00042110 0.2730111 ## Peve_00001263 0.2647003 ## Peve_00034993 0.2618073 ## Peve_00028689 0.2596559 ## Peve_00042617 0.2581788 ## Peve_00033332 0.2440608 ## Peve_00023486 0.2431129 ## Peve_00028711 0.2413697 ## Peve_00004352 0.2355717 ## Peve_00029078 0.2346551 ## Peve_00011114 0.2337310 ## Peve_00004877 0.2326238 ## Peve_00013591 0.2321748 ## Peve_00001380 0.2244332 ## Peve_00022088 0.2226468 ## Peve_00036465 0.2222573 ## Peve_00011010 0.2184097 ## Peve_00001311 0.2182667 ## Peve_00019425 0.2139019 ## Peve_00024470 0.2112225 ## Peve_00033164 0.2061914 ## Peve_00045007 0.2026311 ## Peve_00041730 0.2020993 ## Peve_00001984 0.1988274 ## Peve_00040398 0.1906568 ## Peve_00016236 0.1856058 ## Peve_00020613 0.1824805 ## Peve_00010237 0.1799610 ## Peve_00023599 0.1688783 ## Peve_00006474 0.1622243 ## Peve_00020827 0.1589428 ## Peve_00026219 0.1504053 ## Peve_00022394 0.1443306 ## Peve_00021924 0.1405473 ## Peve_00029077 0.1377499 ## Peve_00042622 0.1368229 ## Peve_00022087 0.1363416 ## Peve_00027038 0.1281876 ## Peve_00026497 0.1239171 ## Peve_00012876 0.1100838 ## Peve_00035220 0.1085293 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1.5) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Lipolysis") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-51-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00018372 1.501795 ## 2 Peve_00009113 1.504824 ## 3 Peve_00044383 1.506173 ## 4 Peve_00012723 1.507918 ## 5 Peve_00030297 1.508158 ## 6 Peve_00024967 1.510926 ## 7 Peve_00042079 1.511757 ## 8 Peve_00010859 1.513388 ## 9 Peve_00026126 1.514429 ## 10 Peve_00017897 1.518210 ## 11 Peve_00030007 1.525757 ## 12 Peve_00037635 1.530104 ## 13 Peve_00031844 1.530114 ## 14 Peve_00044494 1.533230 ## 15 Peve_00000528 1.533676 ## 16 Peve_00024386 1.540863 ## 17 Peve_00001522 1.547200 ## 18 Peve_00003252 1.551435 ## 19 Peve_00013614 1.555154 ## 20 Peve_00011444 1.559656 ## 21 Peve_00043558 1.568027 ## 22 Peve_00033530 1.579383 ## 23 Peve_00028038 1.579649 ## 24 Peve_00031211 1.590058 ## 25 Peve_00034022 1.592555 ## 26 Peve_00022915 1.594099 ## 27 Peve_00024065 1.594517 ## 28 Peve_00021294 1.605998 ## 29 Peve_00002772 1.607455 ## 30 Peve_00024810 1.613918 ## 31 Peve_00032844 1.615793 ## 32 Peve_00006912 1.621844 ## 33 Peve_00023375 1.623123 ## 34 Peve_00019683 1.627228 ## 35 Peve_00016220 1.630525 ## 36 Peve_00034757 1.630860 ## 37 Peve_00002968 1.635887 ## 38 Peve_00000626 1.635979 ## 39 Peve_00030742 1.637783 ## 40 Peve_00017203 1.638150 ## 41 Peve_00030037 1.645170 ## 42 Peve_00006892 1.647543 ## 43 Peve_00026350 1.655750 ## 44 Peve_00028037 1.659583 ## 45 Peve_00008664 1.674231 ## 46 Peve_00001194 1.677296 ## 47 Peve_00015834 1.678868 ## 48 Peve_00006753 1.720037 ## 49 Peve_00004624 1.732280 ## 50 Peve_00027198 1.735028 ## 51 Peve_00038749 1.761132 ## 52 Peve_00036046 1.777657 ## 53 Peve_00016032 1.779320 ## 54 Peve_00032856 1.786050 ## 55 Peve_00021880 1.790300 ## 56 Peve_00022762 1.802784 ## 57 Peve_00036939 1.835560 ## 58 Peve_00037585 1.843671 ## 59 Peve_00036049 1.872458 ## 60 Peve_00023487 1.896950 ## 61 Peve_00001961 1.904562 ## 62 Peve_00038430 1.985082 ## 63 Peve_00005936 1.996480 ## 64 Peve_00033018 2.006105 ## 65 Peve_00032336 2.088259 ## 66 Peve_00021850 2.105384 ## 67 Peve_00006804 2.113135 ## 68 Peve_00017775 2.171518 Plot important genes. ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=Peve_00017775, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-52-1.png) Plot second most important ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=Peve_00006804, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-53-1.png) Plot third most important ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=Peve_00021850, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-54-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 517.49 0.20568 2.9346 0.004 ** ## Residual 34 1998.51 0.79432 ## Total 37 2516.00 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 Significant differences in lipolysis gene expression profile between time points. View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Lipolysis")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-58-1.png) Pull out PC1 score for each sample for GO term. ``` r scores1 <- scaled.pca$x scores1<-as.data.frame(scores1) scores1<-scores1%>%dplyr::select(PC1) scores1$sample<-pca_data_vips$colony_id_corr scores1$timepoint<-pca_data_vips$timepoint scores1<-scores1%>% rename(lipolysis=PC1) scores<-left_join(scores, scores1) ``` ## Joining with `by = join_by(sample, timepoint)` ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis lipolysis ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 # Gene set 4: Fatty acid beta oxidation Load in gene set generated by Apul-energy-go script ``` r ffa_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0006635_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000384 = col_character(), ## `sp|O07610|LCFB_BACSU` = col_character(), ## `27.789` = col_double(), ## `511` = col_double(), ## `323` = col_double(), ## `13` = col_double(), ## `66` = col_double(), ## `561` = col_double(), ## `28` = col_double(), ## `507` = col_double(), ## `8.18e-55` = col_double(), ## `189` = col_double() ## ) Subset gene count matrix for this gene set. ``` r ffa_genes<-Peve_genes%>% filter(rownames(.) %in% ffa_go) ``` Calculate the sum of the total gene set for each sample. ``` r ffa_genes<-as.data.frame(t(ffa_genes)) ffa_genes$Sample<-rownames(ffa_genes) ffa_genes<-ffa_genes %>% rowwise() %>% mutate(ffa_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data4<-left_join(phys, ffa_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data4%>% ggplot(aes(x=timepoint, y=ffa_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-64-1.png) Plot as a PCA. ``` r pca_data <- data4 %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 798.1 0.09894 1.2445 0.194 ## Residual 34 7267.9 0.90106 ## Total 37 8066.0 1.00000 View by timepoint ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=FALSE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Fatty Acid Beta Oxidation")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-68-1.png) Which genes are driving this? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "FFA Beta Oxidation", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-69-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00017775 2.3240019 ## Peve_00006804 2.2455354 ## Peve_00036939 1.9457828 ## Peve_00023487 1.9003601 ## Peve_00032856 1.8394630 ## Peve_00008664 1.7477847 ## Peve_00011444 1.7237536 ## Peve_00003158 1.7226833 ## Peve_00039635 1.6853036 ## Peve_00008132 1.5833793 ## Peve_00018372 1.5602704 ## Peve_00003535 1.5435196 ## Peve_00013614 1.5220648 ## Peve_00041570 1.4989825 ## Peve_00044494 1.4871761 ## Peve_00027083 1.4548913 ## Peve_00039167 1.4401242 ## Peve_00030007 1.4337418 ## Peve_00033476 1.4187384 ## Peve_00035492 1.3994425 ## Peve_00032855 1.3883995 ## Peve_00009162 1.3615691 ## Peve_00021995 1.3496289 ## Peve_00017766 1.3462394 ## Peve_00043472 1.3385807 ## Peve_00014607 1.3378548 ## Peve_00014226 1.3364379 ## Peve_00037697 1.3106115 ## Peve_00024466 1.2926876 ## Peve_00025825 1.2778427 ## Peve_00016361 1.2679170 ## Peve_00010301 1.2434933 ## Peve_00010640 1.2378294 ## Peve_00033189 1.2336998 ## Peve_00028493 1.2231198 ## Peve_00044421 1.2193486 ## Peve_00039903 1.2012316 ## Peve_00033950 1.1963301 ## Peve_00028674 1.1961125 ## Peve_00036060 1.1876943 ## Peve_00034142 1.1671869 ## Peve_00026798 1.1534882 ## Peve_00019287 1.1428545 ## Peve_00028192 1.1399252 ## Peve_00031667 1.1293437 ## Peve_00003361 1.1276770 ## Peve_00011919 1.1215073 ## Peve_00015295 1.1117894 ## Peve_00001876 1.1086602 ## Peve_00014658 1.1078840 ## Peve_00013350 1.1046822 ## Peve_00026567 1.1012700 ## Peve_00029040 1.0974822 ## Peve_00029735 1.0853543 ## Peve_00002423 1.0834566 ## Peve_00001359 1.0832114 ## Peve_00005527 1.0754635 ## Peve_00039634 1.0680830 ## Peve_00014586 1.0680248 ## Peve_00021274 1.0679603 ## Peve_00017927 1.0673329 ## Peve_00041161 1.0668234 ## Peve_00023351 1.0630573 ## Peve_00035298 1.0603940 ## Peve_00018570 1.0595917 ## Peve_00017091 1.0566813 ## Peve_00003219 1.0546425 ## Peve_00029395 1.0534004 ## Peve_00024021 1.0526942 ## Peve_00044990 1.0502196 ## Peve_00008742 1.0462104 ## Peve_00041722 1.0454150 ## Peve_00021861 1.0423000 ## Peve_00017493 1.0359682 ## Peve_00009330 1.0288563 ## Peve_00039448 1.0277233 ## Peve_00028350 1.0243629 ## Peve_00026589 1.0182666 ## Peve_00000568 1.0169526 ## Peve_00042929 1.0149846 ## Peve_00019553 1.0130617 ## Peve_00013503 1.0122846 ## Peve_00042601 1.0095705 ## Peve_00042194 0.9964184 ## Peve_00030402 0.9946561 ## Peve_00023464 0.9945293 ## Peve_00034791 0.9862804 ## Peve_00018687 0.9840791 ## Peve_00032593 0.9818115 ## Peve_00026737 0.9792513 ## Peve_00007754 0.9751108 ## Peve_00007764 0.9748461 ## Peve_00022617 0.9739395 ## Peve_00015302 0.9706532 ## Peve_00021548 0.9689192 ## Peve_00018376 0.9672576 ## Peve_00021042 0.9667425 ## Peve_00033394 0.9563085 ## Peve_00006866 0.9559771 ## Peve_00031823 0.9534237 ## Peve_00020488 0.9502955 ## Peve_00031318 0.9501939 ## Peve_00021461 0.9482097 ## Peve_00002969 0.9408445 ## Peve_00041557 0.9389628 ## Peve_00022156 0.9379819 ## Peve_00041692 0.9377135 ## Peve_00032931 0.9366392 ## Peve_00018374 0.9342473 ## Peve_00020213 0.9342026 ## Peve_00037817 0.9332467 ## Peve_00031444 0.9287995 ## Peve_00041779 0.9216010 ## Peve_00021662 0.9200529 ## Peve_00025870 0.9122628 ## Peve_00036389 0.9063371 ## Peve_00042932 0.9025158 ## Peve_00024469 0.8983325 ## Peve_00003792 0.8882501 ## Peve_00034052 0.8852449 ## Peve_00005055 0.8852115 ## Peve_00002096 0.8837041 ## Peve_00039032 0.8832438 ## Peve_00008763 0.8787634 ## Peve_00043740 0.8768133 ## Peve_00004239 0.8748733 ## Peve_00037724 0.8718898 ## Peve_00029256 0.8689790 ## Peve_00015169 0.8583671 ## Peve_00041483 0.8575959 ## Peve_00034369 0.8572435 ## Peve_00011678 0.8570660 ## Peve_00016543 0.8394219 ## Peve_00008680 0.8391125 ## Peve_00034498 0.8279554 ## Peve_00032651 0.8243689 ## Peve_00043506 0.8241272 ## Peve_00039844 0.8231446 ## Peve_00008231 0.8219558 ## Peve_00008001 0.8135041 ## Peve_00003314 0.8038444 ## Peve_00037709 0.7925431 ## Peve_00038478 0.7869164 ## Peve_00025726 0.7811850 ## Peve_00021976 0.7782692 ## Peve_00039477 0.7702337 ## Peve_00002097 0.7637862 ## Peve_00018373 0.7631705 ## Peve_00014697 0.7589686 ## Peve_00017870 0.7583207 ## Peve_00007821 0.7572040 ## Peve_00012274 0.7568343 ## Peve_00008003 0.7544969 ## Peve_00014701 0.7534921 ## Peve_00029069 0.7424481 ## Peve_00028934 0.7404092 ## Peve_00025258 0.7385960 ## Peve_00017238 0.7365724 ## Peve_00017765 0.7340581 ## Peve_00024585 0.7323089 ## Peve_00034237 0.7300582 ## Peve_00002905 0.7241904 ## Peve_00037434 0.7237990 ## Peve_00033832 0.7218384 ## Peve_00001529 0.7197282 ## Peve_00030254 0.7165321 ## Peve_00005977 0.7131954 ## Peve_00040401 0.6993693 ## Peve_00018354 0.6896263 ## Peve_00015321 0.6865276 ## Peve_00043840 0.6794398 ## Peve_00043014 0.6778110 ## Peve_00003698 0.6739686 ## Peve_00027942 0.6701363 ## Peve_00032682 0.6574362 ## Peve_00010452 0.6562997 ## Peve_00018571 0.6457955 ## Peve_00010728 0.6454230 ## Peve_00033846 0.6357873 ## Peve_00002105 0.6330006 ## Peve_00014227 0.6276167 ## Peve_00000980 0.6206203 ## Peve_00015310 0.6110529 ## Peve_00002248 0.6092868 ## Peve_00019963 0.6071791 ## Peve_00007198 0.6009908 ## Peve_00031192 0.5952094 ## Peve_00030544 0.5929568 ## Peve_00033678 0.5845950 ## Peve_00035682 0.5844242 ## Peve_00029729 0.5839509 ## Peve_00012068 0.5820525 ## Peve_00024115 0.5811102 ## Peve_00020576 0.5758711 ## Peve_00002027 0.5672062 ## Peve_00011920 0.5580918 ## Peve_00002135 0.5478890 ## Peve_00021901 0.5452997 ## Peve_00029227 0.5285517 ## Peve_00006240 0.5275959 ## Peve_00017665 0.5210523 ## Peve_00027663 0.5159669 ## Peve_00008511 0.5013093 ## Peve_00030447 0.4937471 ## Peve_00018045 0.4936202 ## Peve_00027662 0.4630728 ## Peve_00019209 0.4567323 ## Peve_00014920 0.4105430 ## Peve_00021899 0.3825504 ## Peve_00026860 0.3679765 ## Peve_00033569 0.3588673 ## Peve_00017200 0.3528177 ## Peve_00019393 0.3521646 ## Peve_00033229 0.3413434 ## Peve_00034993 0.2853485 ## Peve_00011114 0.2312228 ## Peve_00001311 0.2154329 ## Peve_00013591 0.1833091 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Beta Oxidation") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-70-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00042601 1.009571 ## 2 Peve_00013503 1.012285 ## 3 Peve_00019553 1.013062 ## 4 Peve_00042929 1.014985 ## 5 Peve_00000568 1.016953 ## 6 Peve_00026589 1.018267 ## 7 Peve_00028350 1.024363 ## 8 Peve_00039448 1.027723 ## 9 Peve_00009330 1.028856 ## 10 Peve_00017493 1.035968 ## 11 Peve_00021861 1.042300 ## 12 Peve_00041722 1.045415 ## 13 Peve_00008742 1.046210 ## 14 Peve_00044990 1.050220 ## 15 Peve_00024021 1.052694 ## 16 Peve_00029395 1.053400 ## 17 Peve_00003219 1.054642 ## 18 Peve_00017091 1.056681 ## 19 Peve_00018570 1.059592 ## 20 Peve_00035298 1.060394 ## 21 Peve_00023351 1.063057 ## 22 Peve_00041161 1.066823 ## 23 Peve_00017927 1.067333 ## 24 Peve_00021274 1.067960 ## 25 Peve_00014586 1.068025 ## 26 Peve_00039634 1.068083 ## 27 Peve_00005527 1.075464 ## 28 Peve_00001359 1.083211 ## 29 Peve_00002423 1.083457 ## 30 Peve_00029735 1.085354 ## 31 Peve_00029040 1.097482 ## 32 Peve_00026567 1.101270 ## 33 Peve_00013350 1.104682 ## 34 Peve_00014658 1.107884 ## 35 Peve_00001876 1.108660 ## 36 Peve_00015295 1.111789 ## 37 Peve_00011919 1.121507 ## 38 Peve_00003361 1.127677 ## 39 Peve_00031667 1.129344 ## 40 Peve_00028192 1.139925 ## 41 Peve_00019287 1.142854 ## 42 Peve_00026798 1.153488 ## 43 Peve_00034142 1.167187 ## 44 Peve_00036060 1.187694 ## 45 Peve_00028674 1.196113 ## 46 Peve_00033950 1.196330 ## 47 Peve_00039903 1.201232 ## 48 Peve_00044421 1.219349 ## 49 Peve_00028493 1.223120 ## 50 Peve_00033189 1.233700 ## 51 Peve_00010640 1.237829 ## 52 Peve_00010301 1.243493 ## 53 Peve_00016361 1.267917 ## 54 Peve_00025825 1.277843 ## 55 Peve_00024466 1.292688 ## 56 Peve_00037697 1.310611 ## 57 Peve_00014226 1.336438 ## 58 Peve_00014607 1.337855 ## 59 Peve_00043472 1.338581 ## 60 Peve_00017766 1.346239 ## 61 Peve_00021995 1.349629 ## 62 Peve_00009162 1.361569 ## 63 Peve_00032855 1.388400 ## 64 Peve_00035492 1.399443 ## 65 Peve_00033476 1.418738 ## 66 Peve_00030007 1.433742 ## 67 Peve_00039167 1.440124 ## 68 Peve_00027083 1.454891 ## 69 Peve_00044494 1.487176 ## 70 Peve_00041570 1.498982 ## 71 Peve_00013614 1.522065 ## 72 Peve_00003535 1.543520 ## 73 Peve_00018372 1.560270 ## 74 Peve_00008132 1.583379 ## 75 Peve_00039635 1.685304 ## 76 Peve_00003158 1.722683 ## 77 Peve_00011444 1.723754 ## 78 Peve_00008664 1.747785 ## 79 Peve_00032856 1.839463 ## 80 Peve_00023487 1.900360 ## 81 Peve_00036939 1.945783 ## 82 Peve_00006804 2.245535 ## 83 Peve_00017775 2.324002 Plot important genes. ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=Peve_00017775, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-71-1.png) Plot second most important ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=Peve_00006804, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-72-1.png) Plot third most important ``` r plot<-data3%>% ggplot(aes(x=timepoint, y=Peve_00036939, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-73-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 424.73 0.1383 1.819 0.03 * ## Residual 34 2646.27 0.8617 ## Total 37 3071.00 1.0000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 View by species ``` r plot2<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Fatty acid Beta Oxidation")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot2 ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-77-1.png) Pull out PC1 score for each sample for GO term. ``` r scores1 <- scaled.pca$x scores1<-as.data.frame(scores1) scores1<-scores1%>%dplyr::select(PC1) scores1$sample<-pca_data_vips$colony_id_corr scores1$timepoint<-pca_data_vips$timepoint scores1<-scores1%>% rename(fa_beta=PC1) scores<-left_join(scores, scores1) ``` ## Joining with `by = join_by(sample, timepoint)` ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis lipolysis fa_beta ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 -2.7317639 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 -15.4501240 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 0.2483462 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 7.3058463 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 -12.1525556 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 2.0021942 # Gene set 5: Starvation Load in gene set generated by Apul-energy-go script ``` r starve_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0042594_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000005 = col_character(), ## `sp|Q21307|MEK1_CAEEL` = col_character(), ## `40.134` = col_double(), ## `299` = col_double(), ## `162` = col_double(), ## `7` = col_double(), ## `90` = col_double(), ## `383` = col_double(), ## `34` = col_double(), ## `320` = col_double(), ## `4.31e-69` = col_double(), ## `218` = col_double() ## ) Subset gene count matrix for this gene set. ``` r starve_genes<-Peve_genes%>% filter(rownames(.) %in% starve_go) ``` Calculate the sum of the total gene set for each sample. ``` r starve_genes<-as.data.frame(t(starve_genes)) starve_genes$Sample<-rownames(starve_genes) starve_genes<-starve_genes %>% rowwise() %>% mutate(starve_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data5<-left_join(phys, starve_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data5%>% ggplot(aes(x=timepoint, y=starve_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-83-1.png) Plot as a PCA. ``` r pca_data <- data5 %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 12779 0.11947 1.5377 0.032 * ## Residual 34 94188 0.88053 ## Total 37 106967 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 View by timepoint ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=FALSE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-87-1.png) Which genes are driving this? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "Starvation", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-88-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00018490 2.97596650 ## Peve_00027062 2.20562522 ## Peve_00013660 2.11612585 ## Peve_00020989 2.01845893 ## Peve_00018054 1.99853290 ## Peve_00013900 1.98474546 ## Peve_00036772 1.98208957 ## Peve_00024597 1.96280172 ## Peve_00027223 1.95671770 ## Peve_00011914 1.94567765 ## Peve_00003308 1.93197317 ## Peve_00038749 1.92895438 ## Peve_00015544 1.90244361 ## Peve_00037730 1.90047464 ## Peve_00006912 1.89637526 ## Peve_00016139 1.88907140 ## Peve_00029670 1.88297985 ## Peve_00017775 1.84950953 ## Peve_00042452 1.84935657 ## Peve_00041690 1.84525502 ## Peve_00024410 1.83351395 ## Peve_00021850 1.83188724 ## Peve_00029784 1.82634415 ## Peve_00008454 1.81222098 ## Peve_00009662 1.81049971 ## Peve_00022825 1.80765074 ## Peve_00037415 1.80616899 ## Peve_00015419 1.79214801 ## Peve_00001919 1.76985182 ## Peve_00033018 1.76750519 ## Peve_00017138 1.76702019 ## Peve_00037702 1.76540572 ## Peve_00038744 1.75886414 ## Peve_00034195 1.75453118 ## Peve_00009248 1.75181310 ## Peve_00016713 1.74746252 ## Peve_00011913 1.74631572 ## Peve_00014691 1.74382322 ## Peve_00000174 1.74372727 ## Peve_00007988 1.74188795 ## Peve_00016809 1.74151591 ## Peve_00017929 1.73433176 ## Peve_00034340 1.73219886 ## Peve_00044500 1.72194946 ## Peve_00037277 1.71507937 ## Peve_00012059 1.71153071 ## Peve_00009311 1.70971756 ## Peve_00002619 1.70840852 ## Peve_00027794 1.70395376 ## Peve_00000006 1.70194602 ## Peve_00032944 1.69788450 ## Peve_00037864 1.69756345 ## Peve_00027763 1.69313884 ## Peve_00042557 1.68408851 ## Peve_00013493 1.67469239 ## Peve_00040575 1.67163440 ## Peve_00038468 1.67009080 ## Peve_00029657 1.66341091 ## Peve_00034872 1.66255863 ## Peve_00001961 1.65798413 ## Peve_00045083 1.65659799 ## Peve_00007263 1.65446809 ## Peve_00033939 1.63909934 ## Peve_00001955 1.63849375 ## Peve_00002788 1.63542279 ## Peve_00024967 1.63489590 ## Peve_00032286 1.63048146 ## Peve_00000339 1.63029376 ## Peve_00007065 1.63015883 ## Peve_00043739 1.62835851 ## Peve_00027531 1.62699235 ## Peve_00000494 1.62595049 ## Peve_00021328 1.62336877 ## Peve_00027793 1.62315213 ## Peve_00003830 1.62172882 ## Peve_00001194 1.62133070 ## Peve_00018573 1.61911974 ## Peve_00030742 1.61689422 ## Peve_00021463 1.61686714 ## Peve_00015465 1.61605718 ## Peve_00011121 1.61523879 ## Peve_00038883 1.60871880 ## Peve_00036499 1.60637198 ## Peve_00022762 1.60598927 ## Peve_00033710 1.60526105 ## Peve_00027198 1.60324936 ## Peve_00014560 1.60229955 ## Peve_00015904 1.60222286 ## Peve_00036347 1.59956346 ## Peve_00025358 1.59584104 ## Peve_00035895 1.58764605 ## Peve_00023487 1.58396392 ## Peve_00010859 1.58300715 ## Peve_00025206 1.58054290 ## Peve_00019876 1.57964248 ## Peve_00028567 1.57786571 ## Peve_00033792 1.57461717 ## Peve_00004769 1.57449958 ## Peve_00001586 1.57253887 ## Peve_00028239 1.57160856 ## Peve_00005967 1.56874368 ## Peve_00040036 1.56831737 ## Peve_00019683 1.56519379 ## Peve_00008866 1.56363822 ## Peve_00001903 1.56271724 ## Peve_00008499 1.56226090 ## Peve_00004526 1.55998243 ## Peve_00027250 1.55879383 ## Peve_00006508 1.55705363 ## Peve_00014072 1.55241921 ## Peve_00038504 1.55192919 ## Peve_00023277 1.55149884 ## Peve_00003158 1.55139941 ## Peve_00032881 1.55071709 ## Peve_00034053 1.54861730 ## Peve_00044503 1.54860608 ## Peve_00033992 1.54849413 ## Peve_00040857 1.54667467 ## Peve_00004144 1.54643228 ## Peve_00024602 1.54163132 ## Peve_00023970 1.53654398 ## Peve_00017518 1.53261660 ## Peve_00030194 1.53208106 ## Peve_00021294 1.52888948 ## Peve_00021195 1.52804942 ## Peve_00039635 1.52686390 ## Peve_00014687 1.52601729 ## Peve_00020723 1.52512269 ## Peve_00015416 1.52444031 ## Peve_00029377 1.52443743 ## Peve_00038115 1.52346687 ## Peve_00000528 1.52327740 ## Peve_00013452 1.51851095 ## Peve_00007448 1.51648938 ## Peve_00009007 1.51552599 ## Peve_00042748 1.51382327 ## Peve_00009183 1.50985086 ## Peve_00008524 1.50963053 ## Peve_00044115 1.50835610 ## Peve_00012655 1.50584334 ## Peve_00006523 1.50567555 ## Peve_00023838 1.50264261 ## Peve_00044549 1.50009959 ## Peve_00037424 1.49959900 ## Peve_00018194 1.49834842 ## Peve_00020198 1.49730189 ## Peve_00028072 1.49721302 ## Peve_00011771 1.49705198 ## Peve_00023836 1.49306555 ## Peve_00024527 1.49273405 ## Peve_00017834 1.49016096 ## Peve_00034106 1.48866815 ## Peve_00020417 1.48842818 ## Peve_00013878 1.48816870 ## Peve_00028038 1.48740727 ## Peve_00006859 1.48728349 ## Peve_00033530 1.48617928 ## Peve_00035853 1.48553107 ## Peve_00041076 1.48483049 ## Peve_00026350 1.48299817 ## Peve_00044707 1.48247396 ## Peve_00024600 1.48105501 ## Peve_00017576 1.48063864 ## Peve_00022447 1.47571042 ## Peve_00029795 1.47441387 ## Peve_00006198 1.47360500 ## Peve_00040173 1.47356492 ## Peve_00018044 1.47244052 ## Peve_00004331 1.47187801 ## Peve_00012697 1.47143582 ## Peve_00022326 1.47012131 ## Peve_00014347 1.46831152 ## Peve_00045298 1.46780493 ## Peve_00037732 1.46379526 ## Peve_00029812 1.46276974 ## Peve_00007875 1.46026917 ## Peve_00037078 1.45971431 ## Peve_00036778 1.45836481 ## Peve_00008094 1.45792075 ## Peve_00027249 1.45769608 ## Peve_00007809 1.45708234 ## Peve_00020724 1.45668085 ## Peve_00003252 1.45564922 ## Peve_00034790 1.45538173 ## Peve_00038808 1.45518597 ## Peve_00042048 1.45277765 ## Peve_00039654 1.45229779 ## Peve_00015294 1.45224368 ## Peve_00012025 1.45140249 ## Peve_00006348 1.45078254 ## Peve_00008989 1.44876585 ## Peve_00014804 1.44814410 ## Peve_00018146 1.44809498 ## Peve_00016916 1.44653338 ## Peve_00001478 1.44616063 ## Peve_00019951 1.44483197 ## Peve_00021878 1.44398609 ## Peve_00002527 1.44352059 ## Peve_00016667 1.44274908 ## Peve_00043616 1.44241034 ## Peve_00002968 1.44238982 ## Peve_00013008 1.43927512 ## Peve_00025001 1.43838675 ## Peve_00027168 1.43799460 ## Peve_00029908 1.43790602 ## Peve_00013450 1.43759220 ## Peve_00014579 1.43703109 ## Peve_00023991 1.43576879 ## Peve_00000060 1.43543632 ## Peve_00014941 1.42675165 ## Peve_00028755 1.42601158 ## Peve_00019472 1.42399255 ## Peve_00031895 1.42354715 ## Peve_00043558 1.42234689 ## Peve_00001160 1.42071516 ## Peve_00031211 1.41969376 ## Peve_00014756 1.41932580 ## Peve_00041984 1.41772355 ## Peve_00039642 1.41706840 ## Peve_00042234 1.41699115 ## Peve_00016220 1.41455924 ## Peve_00028481 1.41393085 ## Peve_00026382 1.41283955 ## Peve_00031707 1.41231889 ## Peve_00006545 1.41107766 ## Peve_00014871 1.41074069 ## Peve_00018685 1.40868272 ## Peve_00019033 1.40806249 ## Peve_00044100 1.40778380 ## Peve_00019685 1.40758634 ## Peve_00007645 1.40722281 ## Peve_00018839 1.40487199 ## Peve_00037085 1.40259321 ## Peve_00022915 1.40227071 ## Peve_00020350 1.40214033 ## Peve_00005803 1.40211076 ## Peve_00035887 1.40024624 ## Peve_00040865 1.40017139 ## Peve_00033364 1.39909340 ## Peve_00044383 1.39872300 ## Peve_00029796 1.39789467 ## Peve_00042079 1.39742047 ## Peve_00037558 1.39732295 ## Peve_00024726 1.39479805 ## Peve_00010104 1.39455060 ## Peve_00029340 1.39231022 ## Peve_00003746 1.39194007 ## Peve_00023799 1.39072664 ## Peve_00044994 1.38903805 ## Peve_00009886 1.38784059 ## Peve_00044127 1.38751818 ## Peve_00030037 1.38726348 ## Peve_00039469 1.38592751 ## Peve_00026531 1.38567721 ## Peve_00038456 1.38512017 ## 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Peve_00000517 0.54616253 ## Peve_00024868 0.54581421 ## Peve_00030935 0.54535444 ## Peve_00043854 0.54492034 ## Peve_00003462 0.54480239 ## Peve_00017238 0.54407917 ## Peve_00021564 0.54407693 ## Peve_00007827 0.54398879 ## Peve_00009094 0.54385451 ## Peve_00007162 0.54348019 ## Peve_00023640 0.54107589 ## Peve_00020031 0.53852386 ## Peve_00001304 0.53759444 ## Peve_00033398 0.53713982 ## Peve_00017324 0.53699706 ## Peve_00023576 0.53670316 ## Peve_00023204 0.53668717 ## Peve_00000023 0.53613741 ## Peve_00013221 0.53573476 ## Peve_00006689 0.53567235 ## Peve_00022337 0.53522632 ## Peve_00020567 0.53483333 ## Peve_00024972 0.53452379 ## Peve_00008881 0.53302497 ## Peve_00014797 0.53273229 ## Peve_00007905 0.53270873 ## Peve_00029557 0.53245235 ## Peve_00006976 0.53199120 ## Peve_00026251 0.53192187 ## Peve_00042763 0.53115231 ## Peve_00038234 0.53073260 ## Peve_00018045 0.53069240 ## Peve_00032659 0.52981190 ## Peve_00032543 0.52870985 ## Peve_00017328 0.52867314 ## Peve_00027245 0.52839642 ## Peve_00020719 0.52741547 ## Peve_00004531 0.52679610 ## Peve_00029609 0.52601404 ## Peve_00022976 0.52367365 ## Peve_00022178 0.52268620 ## Peve_00010384 0.52260675 ## Peve_00030861 0.52225494 ## Peve_00037792 0.52214893 ## Peve_00015484 0.52083377 ## Peve_00004071 0.51887553 ## Peve_00025147 0.51850093 ## Peve_00022945 0.51763203 ## Peve_00026860 0.51594321 ## Peve_00009284 0.51543477 ## Peve_00020830 0.51501432 ## Peve_00042684 0.51338731 ## Peve_00018650 0.51191968 ## Peve_00017074 0.51182140 ## Peve_00010723 0.51143749 ## Peve_00008492 0.51142136 ## Peve_00043840 0.51109224 ## Peve_00017294 0.50971587 ## Peve_00027686 0.50925866 ## Peve_00000193 0.50612543 ## Peve_00003037 0.50552108 ## Peve_00001368 0.50519707 ## Peve_00019169 0.50419018 ## Peve_00029983 0.50366991 ## Peve_00013006 0.50272493 ## Peve_00011287 0.50230732 ## Peve_00039650 0.50157326 ## Peve_00035312 0.50017102 ## Peve_00029774 0.49848007 ## Peve_00039845 0.49809397 ## Peve_00004413 0.49789214 ## Peve_00014075 0.49707786 ## Peve_00015292 0.49611490 ## Peve_00018856 0.49555031 ## Peve_00008491 0.49553296 ## Peve_00042791 0.49541831 ## Peve_00018245 0.49421449 ## Peve_00010721 0.49394646 ## Peve_00028722 0.49389430 ## Peve_00041191 0.49382904 ## Peve_00020984 0.49353043 ## Peve_00020571 0.49276143 ## Peve_00018548 0.49268666 ## Peve_00032875 0.49235624 ## Peve_00013177 0.49060791 ## Peve_00012086 0.49038047 ## Peve_00040295 0.48989786 ## Peve_00038116 0.48959306 ## Peve_00006865 0.48946041 ## Peve_00025258 0.48931715 ## Peve_00004587 0.48733123 ## Peve_00004025 0.48675713 ## Peve_00036861 0.48499056 ## Peve_00001721 0.48345910 ## Peve_00016095 0.48305617 ## Peve_00034550 0.48236877 ## Peve_00032368 0.47990636 ## Peve_00034622 0.47974269 ## Peve_00026761 0.47947850 ## Peve_00011581 0.47935432 ## Peve_00027319 0.47931189 ## Peve_00015399 0.47878823 ## Peve_00032498 0.47871630 ## Peve_00042028 0.47816402 ## Peve_00000455 0.47732257 ## Peve_00041559 0.47638525 ## Peve_00008381 0.47469172 ## Peve_00043368 0.47385003 ## Peve_00045210 0.47381824 ## Peve_00023999 0.47336107 ## Peve_00030617 0.47310974 ## Peve_00038665 0.47271083 ## Peve_00040322 0.47260011 ## Peve_00043979 0.47142428 ## Peve_00002061 0.47084491 ## Peve_00027034 0.46940261 ## Peve_00019075 0.46935527 ## Peve_00043672 0.46926532 ## Peve_00039680 0.46842051 ## Peve_00026435 0.46647890 ## Peve_00006860 0.46647548 ## Peve_00005351 0.46566443 ## Peve_00009027 0.46553839 ## Peve_00001233 0.46530192 ## Peve_00043170 0.46506133 ## Peve_00038240 0.46424601 ## Peve_00025174 0.46345020 ## Peve_00004971 0.46297230 ## Peve_00000665 0.46290719 ## Peve_00019170 0.46204920 ## Peve_00018569 0.46076529 ## Peve_00011989 0.45919323 ## Peve_00021875 0.45887360 ## Peve_00024937 0.45856638 ## Peve_00011566 0.45755262 ## Peve_00002586 0.45678988 ## Peve_00030470 0.45652996 ## Peve_00021229 0.45639382 ## Peve_00029545 0.45555363 ## Peve_00002765 0.45454545 ## Peve_00039799 0.45226713 ## Peve_00009266 0.45220861 ## Peve_00031653 0.45204785 ## Peve_00011839 0.45082625 ## Peve_00036030 0.44924879 ## Peve_00025148 0.44912733 ## Peve_00014515 0.44898027 ## Peve_00018127 0.44866656 ## Peve_00033990 0.44800155 ## Peve_00042087 0.44701633 ## Peve_00018491 0.44449275 ## Peve_00042027 0.44383357 ## Peve_00036268 0.43926710 ## Peve_00000715 0.43788655 ## Peve_00040110 0.43684189 ## Peve_00027320 0.43572888 ## Peve_00005211 0.43555700 ## Peve_00016105 0.43463827 ## Peve_00039958 0.43365713 ## Peve_00023032 0.43356664 ## Peve_00024152 0.43318554 ## Peve_00023191 0.43223721 ## Peve_00006977 0.43137756 ## Peve_00004870 0.43095641 ## Peve_00039666 0.42927924 ## Peve_00016103 0.42712168 ## Peve_00032276 0.42632399 ## Peve_00042076 0.42577871 ## Peve_00012937 0.42535623 ## Peve_00023642 0.42523696 ## Peve_00008668 0.42446761 ## Peve_00032757 0.42411398 ## Peve_00032584 0.42375935 ## Peve_00035289 0.42270757 ## Peve_00035828 0.42217438 ## Peve_00016482 0.42199117 ## Peve_00014425 0.42102848 ## Peve_00018390 0.42069229 ## Peve_00018972 0.41976819 ## Peve_00044252 0.41597654 ## Peve_00011775 0.41558811 ## Peve_00017319 0.41441485 ## Peve_00044998 0.41402290 ## Peve_00024055 0.41400216 ## Peve_00015932 0.41309603 ## Peve_00042909 0.41308782 ## Peve_00019239 0.41300834 ## Peve_00018661 0.41026266 ## Peve_00044642 0.41020902 ## Peve_00027976 0.40973985 ## Peve_00028205 0.40807340 ## Peve_00034956 0.40694819 ## Peve_00012123 0.40670869 ## Peve_00035613 0.40501302 ## Peve_00041429 0.40444115 ## Peve_00037889 0.40338359 ## Peve_00035205 0.40316274 ## Peve_00040997 0.40244270 ## Peve_00015519 0.40215004 ## Peve_00028178 0.40116748 ## Peve_00019068 0.40027307 ## Peve_00023484 0.39946458 ## Peve_00037600 0.39812403 ## Peve_00039612 0.39670931 ## Peve_00029076 0.39558065 ## Peve_00024765 0.39453442 ## Peve_00033569 0.39422038 ## Peve_00017861 0.39396493 ## Peve_00039696 0.39377673 ## Peve_00024576 0.39300123 ## Peve_00017854 0.39258494 ## Peve_00017476 0.39223103 ## Peve_00025002 0.39223103 ## Peve_00036890 0.39223103 ## Peve_00038233 0.39223103 ## Peve_00015067 0.39223103 ## Peve_00044948 0.39114988 ## Peve_00004744 0.39045199 ## Peve_00011736 0.38758305 ## Peve_00001131 0.38535715 ## Peve_00000415 0.38234583 ## Peve_00002345 0.38139067 ## Peve_00017883 0.38074329 ## Peve_00002289 0.37836503 ## Peve_00009193 0.37836503 ## Peve_00038168 0.37732512 ## Peve_00037659 0.37643987 ## Peve_00026870 0.37600917 ## Peve_00040637 0.37318563 ## Peve_00030478 0.37288667 ## Peve_00026026 0.36776709 ## Peve_00039319 0.36418110 ## Peve_00034362 0.36371616 ## Peve_00026229 0.36171970 ## Peve_00033636 0.36094493 ## Peve_00024841 0.35863387 ## Peve_00026934 0.35847226 ## Peve_00021899 0.35813700 ## Peve_00015154 0.35775804 ## Peve_00027422 0.35439872 ## Peve_00036162 0.35439872 ## Peve_00029244 0.35384387 ## Peve_00006874 0.35170360 ## Peve_00025473 0.35137679 ## Peve_00000413 0.35108405 ## Peve_00028876 0.34674656 ## Peve_00043438 0.34618090 ## Peve_00014513 0.34571063 ## Peve_00033327 0.34571063 ## Peve_00041688 0.34527901 ## Peve_00016749 0.34491269 ## Peve_00036480 0.34409889 ## Peve_00031651 0.34230555 ## Peve_00035260 0.34060676 ## Peve_00025466 0.33868390 ## Peve_00015061 0.33807467 ## Peve_00001132 0.33692712 ## Peve_00001954 0.33662178 ## Peve_00044754 0.33525491 ## Peve_00022705 0.33508764 ## Peve_00026888 0.33307457 ## Peve_00026771 0.33068651 ## Peve_00040465 0.32983261 ## Peve_00003272 0.32983021 ## Peve_00024083 0.32947824 ## Peve_00013040 0.32899758 ## Peve_00005452 0.32849658 ## Peve_00028643 0.32751624 ## Peve_00024959 0.32310962 ## Peve_00025621 0.32282665 ## Peve_00025528 0.32267677 ## Peve_00044637 0.32029198 ## Peve_00015354 0.31976893 ## Peve_00035220 0.31882476 ## Peve_00039037 0.31761680 ## Peve_00042183 0.31738588 ## Peve_00024296 0.31719646 ## Peve_00037866 0.31659550 ## Peve_00010098 0.31638144 ## Peve_00006862 0.31501907 ## Peve_00032151 0.31092215 ## Peve_00014913 0.31053805 ## Peve_00025095 0.30991518 ## Peve_00033888 0.30912665 ## Peve_00007419 0.30891104 ## Peve_00010237 0.30772420 ## Peve_00020456 0.30717426 ## Peve_00036710 0.30635521 ## Peve_00022290 0.30578643 ## Peve_00034445 0.30485409 ## Peve_00012776 0.30409381 ## Peve_00029677 0.30379837 ## Peve_00040065 0.30153914 ## Peve_00001372 0.30022182 ## Peve_00014523 0.29588647 ## Peve_00021520 0.29322926 ## Peve_00023429 0.29269986 ## Peve_00023800 0.29161525 ## Peve_00014231 0.29023460 ## Peve_00027917 0.28976473 ## Peve_00028036 0.28682285 ## Peve_00005214 0.28348885 ## Peve_00039769 0.28299790 ## Peve_00005448 0.28277518 ## Peve_00027247 0.28241207 ## Peve_00030471 0.28241207 ## Peve_00023299 0.28056583 ## Peve_00020710 0.28055087 ## Peve_00044804 0.27959103 ## Peve_00026254 0.27951437 ## Peve_00016050 0.27613887 ## Peve_00019680 0.27513076 ## Peve_00031596 0.27319938 ## Peve_00008952 0.27282668 ## Peve_00026219 0.27223012 ## Peve_00037417 0.26796959 ## Peve_00042617 0.26622503 ## Peve_00030611 0.26614392 ## Peve_00002344 0.26594264 ## Peve_00011788 0.26431401 ## Peve_00014581 0.26392502 ## Peve_00010507 0.26286739 ## Peve_00012432 0.26153488 ## Peve_00026294 0.25717220 ## Peve_00026773 0.25519580 ## Peve_00027410 0.25147911 ## Peve_00033571 0.25007577 ## Peve_00024500 0.24638864 ## Peve_00012381 0.24330709 ## Peve_00037335 0.24000696 ## Peve_00004877 0.23221409 ## Peve_00007816 0.23085808 ## Peve_00043799 0.23001103 ## Peve_00036455 0.22832383 ## Peve_00007005 0.22744980 ## Peve_00031151 0.22697551 ## Peve_00030571 0.22642321 ## Peve_00032406 0.22634786 ## Peve_00039110 0.22022622 ## Peve_00008694 0.22005070 ## Peve_00021604 0.21748534 ## Peve_00042110 0.21661131 ## Peve_00040489 0.21449802 ## Peve_00024470 0.21395670 ## Peve_00027719 0.21204436 ## Peve_00003982 0.21064378 ## Peve_00032222 0.20981142 ## Peve_00045007 0.20790477 ## Peve_00026871 0.20591059 ## Peve_00029078 0.20343498 ## Peve_00020269 0.20244368 ## Peve_00040546 0.20067410 ## Peve_00017318 0.19938619 ## Peve_00031367 0.19779567 ## Peve_00043853 0.19767940 ## Peve_00001665 0.19755189 ## Peve_00027118 0.19668487 ## Peve_00035172 0.19296151 ## Peve_00014518 0.19200099 ## Peve_00032476 0.19148917 ## Peve_00000179 0.18993855 ## Peve_00036712 0.18512834 ## Peve_00041624 0.18416084 ## Peve_00004633 0.18193191 ## Peve_00038060 0.18161876 ## Peve_00039429 0.18115614 ## Peve_00028711 0.18046285 ## Peve_00042622 0.17494556 ## Peve_00036498 0.17315052 ## Peve_00017488 0.17254449 ## Peve_00020827 0.16190705 ## Peve_00001380 0.16187844 ## Peve_00033894 0.16106443 ## Peve_00028467 0.16098051 ## Peve_00000772 0.15827394 ## Peve_00026925 0.15734692 ## Peve_00006474 0.15573026 ## Peve_00027848 0.14870586 ## Peve_00017370 0.14821929 ## Peve_00020429 0.14527197 ## Peve_00011010 0.14519613 ## Peve_00041730 0.14394194 ## Peve_00025812 0.13865901 ## Peve_00023599 0.13692432 ## Peve_00012641 0.13648706 ## Peve_00022944 0.13437891 ## Peve_00014448 0.13382562 ## Peve_00031555 0.13071169 ## Peve_00035114 0.11130634 ## Peve_00031880 0.10977154 ## Peve_00001733 0.10900984 ## Peve_00040729 0.10182173 ## Peve_00040395 0.09876700 ## Peve_00027035 0.08959598 ## Peve_00043653 0.08772469 ## Peve_00032309 0.08150926 ## Peve_00020849 0.07823810 ## Peve_00042090 0.06813870 ## Peve_00001984 0.05875632 ## Peve_00037077 0.05850915 ## Peve_00044598 0.05358728 ## Peve_00045336 0.04955339 ## Peve_00029077 0.04249267 ## Peve_00036016 0.04220525 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1.5) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Starvation") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-89-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00044549 1.500100 ## 2 Peve_00023838 1.502643 ## 3 Peve_00006523 1.505676 ## 4 Peve_00012655 1.505843 ## 5 Peve_00044115 1.508356 ## 6 Peve_00008524 1.509631 ## 7 Peve_00009183 1.509851 ## 8 Peve_00042748 1.513823 ## 9 Peve_00009007 1.515526 ## 10 Peve_00007448 1.516489 ## 11 Peve_00013452 1.518511 ## 12 Peve_00000528 1.523277 ## 13 Peve_00038115 1.523467 ## 14 Peve_00029377 1.524437 ## 15 Peve_00015416 1.524440 ## 16 Peve_00020723 1.525123 ## 17 Peve_00014687 1.526017 ## 18 Peve_00039635 1.526864 ## 19 Peve_00021195 1.528049 ## 20 Peve_00021294 1.528889 ## 21 Peve_00030194 1.532081 ## 22 Peve_00017518 1.532617 ## 23 Peve_00023970 1.536544 ## 24 Peve_00024602 1.541631 ## 25 Peve_00004144 1.546432 ## 26 Peve_00040857 1.546675 ## 27 Peve_00033992 1.548494 ## 28 Peve_00044503 1.548606 ## 29 Peve_00034053 1.548617 ## 30 Peve_00032881 1.550717 ## 31 Peve_00003158 1.551399 ## 32 Peve_00023277 1.551499 ## 33 Peve_00038504 1.551929 ## 34 Peve_00014072 1.552419 ## 35 Peve_00006508 1.557054 ## 36 Peve_00027250 1.558794 ## 37 Peve_00004526 1.559982 ## 38 Peve_00008499 1.562261 ## 39 Peve_00001903 1.562717 ## 40 Peve_00008866 1.563638 ## 41 Peve_00019683 1.565194 ## 42 Peve_00040036 1.568317 ## 43 Peve_00005967 1.568744 ## 44 Peve_00028239 1.571609 ## 45 Peve_00001586 1.572539 ## 46 Peve_00004769 1.574500 ## 47 Peve_00033792 1.574617 ## 48 Peve_00028567 1.577866 ## 49 Peve_00019876 1.579642 ## 50 Peve_00025206 1.580543 ## 51 Peve_00010859 1.583007 ## 52 Peve_00023487 1.583964 ## 53 Peve_00035895 1.587646 ## 54 Peve_00025358 1.595841 ## 55 Peve_00036347 1.599563 ## 56 Peve_00015904 1.602223 ## 57 Peve_00014560 1.602300 ## 58 Peve_00027198 1.603249 ## 59 Peve_00033710 1.605261 ## 60 Peve_00022762 1.605989 ## 61 Peve_00036499 1.606372 ## 62 Peve_00038883 1.608719 ## 63 Peve_00011121 1.615239 ## 64 Peve_00015465 1.616057 ## 65 Peve_00021463 1.616867 ## 66 Peve_00030742 1.616894 ## 67 Peve_00018573 1.619120 ## 68 Peve_00001194 1.621331 ## 69 Peve_00003830 1.621729 ## 70 Peve_00027793 1.623152 ## 71 Peve_00021328 1.623369 ## 72 Peve_00000494 1.625950 ## 73 Peve_00027531 1.626992 ## 74 Peve_00043739 1.628359 ## 75 Peve_00007065 1.630159 ## 76 Peve_00000339 1.630294 ## 77 Peve_00032286 1.630481 ## 78 Peve_00024967 1.634896 ## 79 Peve_00002788 1.635423 ## 80 Peve_00001955 1.638494 ## 81 Peve_00033939 1.639099 ## 82 Peve_00007263 1.654468 ## 83 Peve_00045083 1.656598 ## 84 Peve_00001961 1.657984 ## 85 Peve_00034872 1.662559 ## 86 Peve_00029657 1.663411 ## 87 Peve_00038468 1.670091 ## 88 Peve_00040575 1.671634 ## 89 Peve_00013493 1.674692 ## 90 Peve_00042557 1.684089 ## 91 Peve_00027763 1.693139 ## 92 Peve_00037864 1.697563 ## 93 Peve_00032944 1.697885 ## 94 Peve_00000006 1.701946 ## 95 Peve_00027794 1.703954 ## 96 Peve_00002619 1.708409 ## 97 Peve_00009311 1.709718 ## 98 Peve_00012059 1.711531 ## 99 Peve_00037277 1.715079 ## 100 Peve_00044500 1.721949 ## 101 Peve_00034340 1.732199 ## 102 Peve_00017929 1.734332 ## 103 Peve_00016809 1.741516 ## 104 Peve_00007988 1.741888 ## 105 Peve_00000174 1.743727 ## 106 Peve_00014691 1.743823 ## 107 Peve_00011913 1.746316 ## 108 Peve_00016713 1.747463 ## 109 Peve_00009248 1.751813 ## 110 Peve_00034195 1.754531 ## 111 Peve_00038744 1.758864 ## 112 Peve_00037702 1.765406 ## 113 Peve_00017138 1.767020 ## 114 Peve_00033018 1.767505 ## 115 Peve_00001919 1.769852 ## 116 Peve_00015419 1.792148 ## 117 Peve_00037415 1.806169 ## 118 Peve_00022825 1.807651 ## 119 Peve_00009662 1.810500 ## 120 Peve_00008454 1.812221 ## 121 Peve_00029784 1.826344 ## 122 Peve_00021850 1.831887 ## 123 Peve_00024410 1.833514 ## 124 Peve_00041690 1.845255 ## 125 Peve_00042452 1.849357 ## 126 Peve_00017775 1.849510 ## 127 Peve_00029670 1.882980 ## 128 Peve_00016139 1.889071 ## 129 Peve_00006912 1.896375 ## 130 Peve_00037730 1.900475 ## 131 Peve_00015544 1.902444 ## 132 Peve_00038749 1.928954 ## 133 Peve_00003308 1.931973 ## 134 Peve_00011914 1.945678 ## 135 Peve_00027223 1.956718 ## 136 Peve_00024597 1.962802 ## 137 Peve_00036772 1.982090 ## 138 Peve_00013900 1.984745 ## 139 Peve_00018054 1.998533 ## 140 Peve_00020989 2.018459 ## 141 Peve_00013660 2.116126 ## 142 Peve_00027062 2.205625 ## 143 Peve_00018490 2.975966 Plot important genes. ``` r plot<-data5%>% ggplot(aes(x=timepoint, y=Peve_00018490, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-90-1.png) Plot second most important ``` r plot<-data5%>% ggplot(aes(x=timepoint, y=Peve_00027062, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-91-1.png) Plot third most important ``` r plot<-data5%>% ggplot(aes(x=timepoint, y=Peve_00013660, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-92-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 1261.4 0.2384 3.5477 0.001 *** ## Residual 34 4029.6 0.7616 ## Total 37 5291.0 1.0000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 Significant differences in fatty acid beta oxidation gene expression profile between time points. View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Starvation")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-96-1.png) Pull out PC1 score for each sample for GO term. ``` r scores1 <- scaled.pca$x scores1<-as.data.frame(scores1) scores1<-scores1%>%dplyr::select(PC1) scores1$sample<-pca_data_vips$colony_id_corr scores1$timepoint<-pca_data_vips$timepoint scores1<-scores1%>% rename(starve=PC1) scores<-left_join(scores, scores1) ``` ## Joining with `by = join_by(sample, timepoint)` ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis lipolysis fa_beta starve ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 -2.7317639 -17.737096 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 -15.4501240 -4.815820 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 0.2483462 4.222578 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 7.3058463 6.012566 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 -12.1525556 -3.705685 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 2.0021942 7.931360 # Gene set 6: Lipid biosynthesis Load in gene set generated by Apul-energy-go script ``` r lipid_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0008610_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000047 = col_character(), ## `sp|P22570|ADRO_HUMAN` = col_character(), ## `56.329` = col_double(), ## `474` = col_double(), ## `185` = col_double(), ## `5` = col_double(), ## `40` = col_double(), ## `508` = col_double(), ## `32` = col_double(), ## `488` = col_double(), ## `0.0` = col_double(), ## `522` = col_double() ## ) Subset gene count matrix for this gene set. ``` r lipid_genes<-Peve_genes%>% filter(rownames(.) %in% lipid_go) ``` Calculate the sum of the total gene set for each sample. ``` r lipid_genes<-as.data.frame(t(lipid_genes)) lipid_genes$Sample<-rownames(lipid_genes) lipid_genes<-lipid_genes %>% rowwise() %>% mutate(lipid_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data6<-left_join(phys, lipid_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data6%>% ggplot(aes(x=timepoint, y=lipid_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-102-1.png) Plot as a PCA. ``` r pca_data <- data6 %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 8787 0.11005 1.4015 0.082 . ## Residual 34 71059 0.88995 ## Total 37 79846 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 Significant differences in ffa beta oxidation gene expression profile between time points. View by timepoint ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=FALSE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-106-1.png) Which genes are driving this? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "Lipid Synthesis", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-107-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00018490 2.67575231 ## Peve_00034699 2.00156276 ## Peve_00002400 1.96975247 ## Peve_00042780 1.95798740 ## Peve_00017775 1.94025261 ## Peve_00022347 1.91003521 ## Peve_00008454 1.89455543 ## Peve_00029784 1.89188663 ## Peve_00015419 1.87629388 ## Peve_00038744 1.87165513 ## Peve_00037730 1.86970460 ## Peve_00042452 1.86927988 ## Peve_00034356 1.85886639 ## Peve_00017536 1.84415881 ## Peve_00016038 1.83681542 ## Peve_00011873 1.81034000 ## Peve_00032944 1.80835748 ## Peve_00034195 1.80650079 ## Peve_00013023 1.79549776 ## Peve_00043069 1.79341935 ## Peve_00011914 1.79047304 ## Peve_00014691 1.79007452 ## Peve_00009355 1.78829400 ## Peve_00009248 1.78719221 ## Peve_00008542 1.78646636 ## Peve_00037277 1.76416825 ## Peve_00018054 1.76329482 ## Peve_00033226 1.74499676 ## Peve_00040575 1.74093338 ## Peve_00028225 1.73289588 ## Peve_00014510 1.72636880 ## Peve_00031465 1.71709136 ## Peve_00002788 1.71708852 ## Peve_00043297 1.70761109 ## Peve_00015544 1.70666673 ## Peve_00038749 1.70242303 ## Peve_00039206 1.69782787 ## Peve_00020146 1.69377425 ## Peve_00043739 1.68404247 ## Peve_00036347 1.67195827 ## Peve_00025206 1.66973138 ## Peve_00038883 1.66877471 ## Peve_00011105 1.66664924 ## Peve_00037864 1.66350404 ## Peve_00006912 1.66204807 ## Peve_00024407 1.66181638 ## Peve_00037743 1.65877483 ## Peve_00013024 1.65433844 ## Peve_00013568 1.65221530 ## Peve_00016713 1.64494852 ## Peve_00010928 1.64163400 ## Peve_00003830 1.63445520 ## Peve_00004144 1.63260286 ## Peve_00034365 1.62711060 ## Peve_00006753 1.62448894 ## Peve_00027250 1.61715521 ## Peve_00011913 1.61680896 ## Peve_00016032 1.61648331 ## Peve_00011217 1.61615618 ## Peve_00011918 1.61178851 ## Peve_00042754 1.61156892 ## Peve_00010343 1.59771397 ## Peve_00018194 1.59578596 ## Peve_00004765 1.59063071 ## Peve_00024563 1.58946846 ## Peve_00038287 1.58727837 ## Peve_00041491 1.58668038 ## Peve_00019891 1.58411871 ## Peve_00032856 1.58064476 ## Peve_00022762 1.58055695 ## Peve_00014687 1.58018732 ## Peve_00020147 1.57899705 ## Peve_00038295 1.57769736 ## Peve_00006523 1.57639403 ## Peve_00037702 1.57566964 ## Peve_00019272 1.57486911 ## Peve_00028037 1.57428266 ## Peve_00001739 1.57407422 ## Peve_00030284 1.57404851 ## Peve_00009109 1.56802773 ## Peve_00017834 1.56664533 ## Peve_00039622 1.56483075 ## Peve_00034329 1.56026295 ## Peve_00041076 1.55308037 ## Peve_00015904 1.55120325 ## Peve_00030194 1.55111150 ## Peve_00005967 1.55104688 ## Peve_00029424 1.54954687 ## Peve_00002619 1.54383314 ## Peve_00004624 1.53518134 ## Peve_00043039 1.52981027 ## Peve_00037078 1.52788301 ## Peve_00042598 1.52714506 ## Peve_00044826 1.52713867 ## Peve_00007809 1.52673640 ## Peve_00012025 1.52427633 ## Peve_00016916 1.52334088 ## Peve_00023763 1.52154491 ## Peve_00040173 1.52034621 ## Peve_00023991 1.52002307 ## Peve_00016667 1.51965515 ## Peve_00008664 1.51637810 ## Peve_00039725 1.51516306 ## Peve_00014756 1.51451088 ## Peve_00003702 1.51410446 ## Peve_00030742 1.51373962 ## Peve_00019273 1.51066193 ## Peve_00030037 1.51042886 ## Peve_00041984 1.50995689 ## Peve_00043964 1.50492447 ## Peve_00026069 1.50225081 ## Peve_00018685 1.50042482 ## Peve_00032390 1.49871259 ## Peve_00002772 1.49862987 ## Peve_00035887 1.49797719 ## Peve_00017580 1.49609930 ## Peve_00006892 1.49226317 ## Peve_00043514 1.49211509 ## Peve_00041457 1.49197690 ## Peve_00004790 1.48990698 ## Peve_00020723 1.48891406 ## Peve_00001522 1.48798664 ## Peve_00021878 1.48608846 ## Peve_00032881 1.48508743 ## Peve_00017147 1.47909290 ## Peve_00031407 1.47461043 ## Peve_00025189 1.47211278 ## Peve_00012956 1.47170914 ## Peve_00032709 1.47118495 ## Peve_00001617 1.46929704 ## Peve_00037558 1.46894921 ## Peve_00016220 1.46762674 ## Peve_00006247 1.46602785 ## Peve_00008114 1.46541924 ## Peve_00000545 1.46518640 ## Peve_00022636 1.46103788 ## Peve_00030007 1.46103287 ## Peve_00008302 1.45947075 ## Peve_00018055 1.45540598 ## Peve_00015767 1.45448970 ## Peve_00014665 1.45154149 ## Peve_00008866 1.45104503 ## Peve_00032986 1.44998464 ## Peve_00002031 1.44962664 ## Peve_00015220 1.44820444 ## Peve_00026316 1.44647428 ## Peve_00006508 1.44570816 ## Peve_00036042 1.44514454 ## Peve_00008094 1.44471672 ## Peve_00028786 1.44379978 ## Peve_00025493 1.44345684 ## Peve_00010859 1.44238898 ## Peve_00028254 1.43990087 ## Peve_00025642 1.43988813 ## Peve_00009886 1.43920047 ## Peve_00034757 1.43874982 ## Peve_00034331 1.43314807 ## Peve_00020724 1.43235549 ## Peve_00039207 1.43073801 ## Peve_00017291 1.42672477 ## Peve_00038427 1.42635893 ## Peve_00038456 1.42496545 ## Peve_00028038 1.42473847 ## Peve_00039561 1.42429877 ## Peve_00001366 1.42181563 ## Peve_00000626 1.42161069 ## Peve_00016030 1.41941242 ## Peve_00003076 1.41939047 ## Peve_00003746 1.41771836 ## Peve_00037375 1.41690700 ## Peve_00026920 1.41500324 ## Peve_00031072 1.41373800 ## Peve_00024810 1.41105346 ## Peve_00033934 1.41093879 ## Peve_00009537 1.40998089 ## Peve_00045298 1.40778867 ## Peve_00026126 1.40704364 ## Peve_00034022 1.40572806 ## Peve_00000818 1.40110814 ## Peve_00044181 1.39720875 ## Peve_00023838 1.39663025 ## Peve_00025743 1.39616048 ## Peve_00025176 1.39528852 ## Peve_00016771 1.39492751 ## Peve_00027210 1.39240484 ## Peve_00032844 1.39214277 ## Peve_00031366 1.39182413 ## Peve_00024572 1.39146152 ## Peve_00002328 1.39080730 ## Peve_00021294 1.39035055 ## Peve_00007834 1.38991719 ## Peve_00018947 1.38869487 ## Peve_00009605 1.38830349 ## Peve_00000638 1.38776701 ## Peve_00007801 1.38731701 ## Peve_00002594 1.38721378 ## Peve_00007133 1.38699710 ## Peve_00038604 1.38540081 ## Peve_00028070 1.38474495 ## Peve_00008570 1.38422695 ## Peve_00007412 1.38421634 ## Peve_00010088 1.38187722 ## Peve_00043616 1.38175734 ## Peve_00032737 1.38125613 ## Peve_00012936 1.38118761 ## Peve_00000061 1.38067155 ## Peve_00011525 1.38043494 ## Peve_00010377 1.37962482 ## Peve_00026040 1.37540720 ## Peve_00031773 1.37473126 ## Peve_00038504 1.37422511 ## Peve_00021992 1.37345518 ## Peve_00042522 1.37261548 ## Peve_00037144 1.37234200 ## Peve_00035290 1.37213150 ## Peve_00036567 1.37159444 ## Peve_00000404 1.37125941 ## Peve_00031489 1.37089457 ## Peve_00024829 1.37038519 ## Peve_00033618 1.36907343 ## Peve_00034700 1.36748077 ## Peve_00041570 1.36634114 ## Peve_00006537 1.36564082 ## Peve_00036103 1.36520600 ## Peve_00032879 1.36392751 ## Peve_00037483 1.36217630 ## Peve_00044494 1.36147907 ## Peve_00001843 1.36043091 ## Peve_00044324 1.36028565 ## Peve_00033597 1.36005611 ## Peve_00009182 1.35994585 ## Peve_00040655 1.35927461 ## Peve_00017766 1.35757736 ## Peve_00028939 1.35580995 ## Peve_00019287 1.35253955 ## Peve_00011698 1.35146019 ## Peve_00044288 1.35042375 ## Peve_00043663 1.34900255 ## Peve_00024123 1.34850083 ## Peve_00033983 1.34776934 ## Peve_00031923 1.34720702 ## Peve_00037732 1.34657839 ## Peve_00024075 1.34535405 ## Peve_00026519 1.34439654 ## Peve_00024567 1.34401948 ## Peve_00016176 1.34299230 ## Peve_00042079 1.34100212 ## Peve_00013174 1.34076178 ## Peve_00029137 1.33994971 ## Peve_00033117 1.33985062 ## Peve_00014692 1.33748984 ## Peve_00036458 1.33691001 ## Peve_00033605 1.33183986 ## Peve_00037892 1.33154248 ## Peve_00027163 1.33129558 ## Peve_00021483 1.33129354 ## Peve_00004480 1.33106931 ## Peve_00028239 1.33042265 ## Peve_00003309 1.33015358 ## Peve_00028558 1.33013870 ## Peve_00045032 1.32944532 ## Peve_00007013 1.32783584 ## Peve_00000296 1.32650470 ## Peve_00015148 1.32595302 ## Peve_00024010 1.32528240 ## Peve_00003525 1.32519182 ## Peve_00034363 1.32479473 ## Peve_00019033 1.32460326 ## Peve_00042048 1.32432258 ## Peve_00043013 1.32430526 ## Peve_00010858 1.32351381 ## Peve_00037800 1.32322887 ## Peve_00042234 1.32157894 ## Peve_00031897 1.32131617 ## Peve_00006989 1.32121650 ## Peve_00012251 1.32052188 ## Peve_00007881 1.32000877 ## Peve_00027164 1.31945308 ## Peve_00035492 1.31860628 ## Peve_00029814 1.31735867 ## Peve_00029645 1.31636535 ## Peve_00013214 1.31310824 ## Peve_00039654 1.31280373 ## Peve_00014996 1.31168432 ## Peve_00015247 1.30980401 ## Peve_00039167 1.30966246 ## Peve_00032987 1.30939604 ## Peve_00020048 1.30859117 ## Peve_00024280 1.30855912 ## Peve_00007931 1.30729490 ## Peve_00012877 1.30636017 ## Peve_00033364 1.30555591 ## Peve_00006971 1.30548868 ## Peve_00039469 1.30540344 ## Peve_00012136 1.30539080 ## Peve_00027087 1.30411148 ## Peve_00005770 1.30376989 ## Peve_00034484 1.30342243 ## Peve_00008533 1.30312125 ## Peve_00011789 1.30028851 ## Peve_00018719 1.29944052 ## Peve_00026382 1.29778407 ## Peve_00044487 1.29765620 ## Peve_00001929 1.29608290 ## Peve_00038751 1.29420351 ## Peve_00023836 1.29140200 ## Peve_00028851 1.28972563 ## Peve_00019286 1.28935724 ## Peve_00032474 1.28870241 ## Peve_00043557 1.28833776 ## Peve_00038767 1.28790276 ## Peve_00044059 1.28739398 ## Peve_00030516 1.28677826 ## Peve_00002221 1.28662597 ## Peve_00001262 1.28613545 ## Peve_00013822 1.28591498 ## Peve_00033476 1.28589771 ## Peve_00017204 1.28577891 ## Peve_00032648 1.28559561 ## Peve_00033084 1.28526933 ## Peve_00028437 1.28506964 ## Peve_00009162 1.28485120 ## Peve_00023210 1.28317872 ## Peve_00037482 1.28303455 ## Peve_00016917 1.28122785 ## Peve_00013958 1.28118072 ## Peve_00023491 1.28113538 ## Peve_00039635 1.28087669 ## Peve_00021288 1.28035027 ## Peve_00001870 1.27990389 ## Peve_00030219 1.27963157 ## Peve_00042077 1.27924798 ## Peve_00032855 1.27773713 ## Peve_00032914 1.27746971 ## Peve_00016101 1.27681454 ## Peve_00028559 1.27666468 ## Peve_00033589 1.27581851 ## Peve_00033242 1.27563287 ## Peve_00004158 1.27464208 ## Peve_00000817 1.27356018 ## Peve_00032301 1.27328257 ## Peve_00017913 1.27286016 ## Peve_00019727 1.27189742 ## Peve_00006776 1.27167124 ## Peve_00021477 1.27122358 ## Peve_00010104 1.27056128 ## Peve_00041684 1.27025368 ## Peve_00020889 1.27007219 ## Peve_00004715 1.26998112 ## Peve_00017897 1.26973173 ## Peve_00023334 1.26885080 ## Peve_00033950 1.26848577 ## Peve_00030758 1.26829520 ## Peve_00024030 1.26803309 ## Peve_00018205 1.26740099 ## Peve_00025408 1.26653374 ## Peve_00022381 1.26630616 ## Peve_00017334 1.26567356 ## Peve_00039499 1.26564626 ## Peve_00038841 1.26531887 ## Peve_00044743 1.26526357 ## Peve_00010647 1.26460720 ## Peve_00009666 1.26387948 ## Peve_00003252 1.26371946 ## Peve_00042552 1.26369753 ## Peve_00022075 1.26306823 ## Peve_00001931 1.26301651 ## Peve_00021243 1.26162520 ## Peve_00012294 1.26134718 ## Peve_00037340 1.26102910 ## Peve_00034684 1.26092909 ## Peve_00029507 1.26009608 ## Peve_00036559 1.25809408 ## Peve_00024022 1.25769053 ## Peve_00017958 1.25641768 ## Peve_00043499 1.25539563 ## Peve_00005364 1.25503099 ## Peve_00008973 1.25472093 ## Peve_00018858 1.25464684 ## Peve_00010982 1.25411876 ## Peve_00034013 1.25372040 ## Peve_00017721 1.25226132 ## Peve_00014875 1.25174797 ## Peve_00019983 1.24924067 ## Peve_00016511 1.24849594 ## Peve_00005940 1.24512620 ## Peve_00014689 1.24405072 ## Peve_00026750 1.24337903 ## Peve_00013435 1.24254056 ## Peve_00040903 1.24179797 ## Peve_00025628 1.24132539 ## Peve_00044122 1.24051911 ## Peve_00014021 1.24049147 ## Peve_00003623 1.23979134 ## Peve_00014264 1.23956509 ## Peve_00034187 1.23835913 ## Peve_00042017 1.23805015 ## Peve_00011611 1.23773691 ## Peve_00014814 1.23767160 ## 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Peve_00003272 0.34036603 ## Peve_00031651 0.33857193 ## Peve_00014913 0.33653034 ## Peve_00031478 0.33566536 ## Peve_00017308 0.33527529 ## Peve_00044754 0.33286487 ## Peve_00033771 0.33244774 ## Peve_00003323 0.33188859 ## Peve_00002058 0.32952977 ## Peve_00017306 0.32799972 ## Peve_00033636 0.32624448 ## Peve_00026773 0.32536226 ## Peve_00026134 0.32293197 ## Peve_00031997 0.32013870 ## Peve_00020699 0.31675013 ## Peve_00013786 0.31544066 ## Peve_00026319 0.31337305 ## Peve_00022290 0.31099485 ## Peve_00002289 0.30992865 ## Peve_00031941 0.30821608 ## Peve_00035220 0.30807883 ## Peve_00009152 0.30621053 ## Peve_00015354 0.30501620 ## Peve_00015067 0.30326332 ## Peve_00029929 0.30251539 ## Peve_00011114 0.29839508 ## Peve_00007290 0.29796499 ## Peve_00020613 0.28932434 ## Peve_00022705 0.28824879 ## Peve_00024842 0.28700686 ## Peve_00010095 0.28682700 ## Peve_00029671 0.28276101 ## Peve_00036162 0.28088005 ## Peve_00025621 0.28023705 ## Peve_00028036 0.27813080 ## Peve_00005214 0.27754558 ## Peve_00012779 0.27719160 ## Peve_00012432 0.27607825 ## Peve_00001458 0.27008501 ## Peve_00030810 0.25788187 ## Peve_00044804 0.25591111 ## Peve_00031932 0.25407585 ## Peve_00034445 0.25390829 ## Peve_00036913 0.25025512 ## Peve_00003982 0.24679515 ## Peve_00014770 0.24677271 ## Peve_00022881 0.24620267 ## Peve_00037335 0.24457606 ## Peve_00025131 0.24435877 ## Peve_00043853 0.23619613 ## Peve_00032644 0.23517585 ## Peve_00040704 0.23295735 ## Peve_00031880 0.23227953 ## Peve_00035858 0.23220112 ## Peve_00021608 0.22944374 ## Peve_00035871 0.22806259 ## Peve_00026273 0.22738731 ## Peve_00014506 0.22516018 ## Peve_00041445 0.22095457 ## Peve_00007901 0.21812466 ## Peve_00014460 0.21594683 ## Peve_00027038 0.21460200 ## Peve_00040398 0.21454518 ## Peve_00033164 0.21430571 ## Peve_00001311 0.21193117 ## Peve_00019535 0.20753299 ## Peve_00009264 0.20570212 ## Peve_00042110 0.20441062 ## Peve_00040636 0.20230653 ## Peve_00028689 0.19489428 ## Peve_00006474 0.19428212 ## Peve_00028671 0.19281608 ## Peve_00034366 0.19206240 ## Peve_00045078 0.18979128 ## Peve_00001984 0.18521842 ## Peve_00040652 0.18457346 ## Peve_00026771 0.18446760 ## Peve_00002121 0.18089953 ## Peve_00019425 0.18016038 ## Peve_00012776 0.17744654 ## Peve_00016236 0.17462775 ## Peve_00007816 0.16557846 ## Peve_00013897 0.16277177 ## Peve_00011528 0.16193567 ## Peve_00000772 0.15887383 ## Peve_00026925 0.15481259 ## Peve_00017488 0.15467303 ## Peve_00033894 0.14186906 ## Peve_00017370 0.14049429 ## Peve_00031111 0.13898059 ## Peve_00031555 0.13667510 ## Peve_00017307 0.13188656 ## Peve_00019827 0.11581748 ## Peve_00025622 0.11380766 ## Peve_00027035 0.10933811 ## Peve_00014772 0.10275063 ## Peve_00012876 0.10219852 ## Peve_00012641 0.10108263 ## Peve_00042090 0.09497354 ## Peve_00045336 0.08089905 ## Peve_00019750 0.07594955 ## Peve_00043653 0.02366452 ## Peve_00037077 0.02175467 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1.5) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Lipid Catabolism") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-108-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00018685 1.500425 ## 2 Peve_00026069 1.502251 ## 3 Peve_00043964 1.504924 ## 4 Peve_00041984 1.509957 ## 5 Peve_00030037 1.510429 ## 6 Peve_00019273 1.510662 ## 7 Peve_00030742 1.513740 ## 8 Peve_00003702 1.514104 ## 9 Peve_00014756 1.514511 ## 10 Peve_00039725 1.515163 ## 11 Peve_00008664 1.516378 ## 12 Peve_00016667 1.519655 ## 13 Peve_00023991 1.520023 ## 14 Peve_00040173 1.520346 ## 15 Peve_00023763 1.521545 ## 16 Peve_00016916 1.523341 ## 17 Peve_00012025 1.524276 ## 18 Peve_00007809 1.526736 ## 19 Peve_00044826 1.527139 ## 20 Peve_00042598 1.527145 ## 21 Peve_00037078 1.527883 ## 22 Peve_00043039 1.529810 ## 23 Peve_00004624 1.535181 ## 24 Peve_00002619 1.543833 ## 25 Peve_00029424 1.549547 ## 26 Peve_00005967 1.551047 ## 27 Peve_00030194 1.551112 ## 28 Peve_00015904 1.551203 ## 29 Peve_00041076 1.553080 ## 30 Peve_00034329 1.560263 ## 31 Peve_00039622 1.564831 ## 32 Peve_00017834 1.566645 ## 33 Peve_00009109 1.568028 ## 34 Peve_00030284 1.574049 ## 35 Peve_00001739 1.574074 ## 36 Peve_00028037 1.574283 ## 37 Peve_00019272 1.574869 ## 38 Peve_00037702 1.575670 ## 39 Peve_00006523 1.576394 ## 40 Peve_00038295 1.577697 ## 41 Peve_00020147 1.578997 ## 42 Peve_00014687 1.580187 ## 43 Peve_00022762 1.580557 ## 44 Peve_00032856 1.580645 ## 45 Peve_00019891 1.584119 ## 46 Peve_00041491 1.586680 ## 47 Peve_00038287 1.587278 ## 48 Peve_00024563 1.589468 ## 49 Peve_00004765 1.590631 ## 50 Peve_00018194 1.595786 ## 51 Peve_00010343 1.597714 ## 52 Peve_00042754 1.611569 ## 53 Peve_00011918 1.611789 ## 54 Peve_00011217 1.616156 ## 55 Peve_00016032 1.616483 ## 56 Peve_00011913 1.616809 ## 57 Peve_00027250 1.617155 ## 58 Peve_00006753 1.624489 ## 59 Peve_00034365 1.627111 ## 60 Peve_00004144 1.632603 ## 61 Peve_00003830 1.634455 ## 62 Peve_00010928 1.641634 ## 63 Peve_00016713 1.644949 ## 64 Peve_00013568 1.652215 ## 65 Peve_00013024 1.654338 ## 66 Peve_00037743 1.658775 ## 67 Peve_00024407 1.661816 ## 68 Peve_00006912 1.662048 ## 69 Peve_00037864 1.663504 ## 70 Peve_00011105 1.666649 ## 71 Peve_00038883 1.668775 ## 72 Peve_00025206 1.669731 ## 73 Peve_00036347 1.671958 ## 74 Peve_00043739 1.684042 ## 75 Peve_00020146 1.693774 ## 76 Peve_00039206 1.697828 ## 77 Peve_00038749 1.702423 ## 78 Peve_00015544 1.706667 ## 79 Peve_00043297 1.707611 ## 80 Peve_00002788 1.717089 ## 81 Peve_00031465 1.717091 ## 82 Peve_00014510 1.726369 ## 83 Peve_00028225 1.732896 ## 84 Peve_00040575 1.740933 ## 85 Peve_00033226 1.744997 ## 86 Peve_00018054 1.763295 ## 87 Peve_00037277 1.764168 ## 88 Peve_00008542 1.786466 ## 89 Peve_00009248 1.787192 ## 90 Peve_00009355 1.788294 ## 91 Peve_00014691 1.790075 ## 92 Peve_00011914 1.790473 ## 93 Peve_00043069 1.793419 ## 94 Peve_00013023 1.795498 ## 95 Peve_00034195 1.806501 ## 96 Peve_00032944 1.808357 ## 97 Peve_00011873 1.810340 ## 98 Peve_00016038 1.836815 ## 99 Peve_00017536 1.844159 ## 100 Peve_00034356 1.858866 ## 101 Peve_00042452 1.869280 ## 102 Peve_00037730 1.869705 ## 103 Peve_00038744 1.871655 ## 104 Peve_00015419 1.876294 ## 105 Peve_00029784 1.891887 ## 106 Peve_00008454 1.894555 ## 107 Peve_00022347 1.910035 ## 108 Peve_00017775 1.940253 ## 109 Peve_00042780 1.957987 ## 110 Peve_00002400 1.969752 ## 111 Peve_00034699 2.001563 ## 112 Peve_00018490 2.675752 Plot important genes. ``` r plot<-data6%>% ggplot(aes(x=timepoint, y=Peve_00018490, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-109-1.png) Plot second most important ``` r plot<-data6%>% ggplot(aes(x=timepoint, y=Peve_00034699, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-110-1.png) Plot third most important ``` r plot<-data6%>% ggplot(aes(x=timepoint, y=Peve_00002400, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-111-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 1032.3 0.2491 3.7596 0.001 *** ## Residual 34 3111.7 0.7509 ## Total 37 4144.0 1.0000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 Significant differences in lipid synthesis gene expression profile between time points. View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Lipid Synthesis")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-115-1.png) Pull out PC1 score for each sample for GO term. ``` r scores1 <- scaled.pca$x scores1<-as.data.frame(scores1) scores1<-scores1%>%dplyr::select(PC1) scores1$sample<-pca_data_vips$colony_id_corr scores1$timepoint<-pca_data_vips$timepoint scores1<-scores1%>% rename(lipids=PC1) scores<-left_join(scores, scores1) ``` ## Joining with `by = join_by(sample, timepoint)` ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis lipolysis fa_beta starve ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 -2.7317639 -17.737096 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 -15.4501240 -4.815820 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 0.2483462 4.222578 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 7.3058463 6.012566 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 -12.1525556 -3.705685 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 2.0021942 7.931360 ## lipids ## 1 -16.022543 ## 2 -3.371010 ## 3 4.226094 ## 4 5.368042 ## 5 -3.538633 ## 6 7.235845 # Gene set 7: Protein catabolic process Load in gene set generated by Apul-energy-go script ``` r protein_go<-read_table(file="E-Peve/output/05-Peve-energy-GO/Apul_blastp-GO:0030163_out.tab")%>%pull(var=1) ``` ## ## ── Column specification ──────────────────────────────────────────────────────── ## cols( ## Peve_00000002 = col_character(), ## `sp|B8NSV5|CREB_ASPFN` = col_character(), ## `30.451` = col_double(), ## `266` = col_double(), ## `138` = col_double(), ## `7` = col_double(), ## `185` = col_double(), ## `438` = col_double(), ## `196` = col_double(), ## `426` = col_double(), ## `4.09e-23` = col_double(), ## `102` = col_double() ## ) Subset gene count matrix for this gene set. ``` r protein_genes<-Peve_genes%>% filter(rownames(.) %in% protein_go) ``` Calculate the sum of the total gene set for each sample. ``` r protein_genes<-as.data.frame(t(protein_genes)) protein_genes$Sample<-rownames(protein_genes) protein_genes<-protein_genes %>% rowwise() %>% mutate(protein_count = sum(c_across(where(is.numeric)))) %>% ungroup()%>% as.data.frame() ``` Merge into master data frame with metadata and physiology as a new column called “glycolysis”. ``` r data7<-left_join(phys, protein_genes) ``` ## Joining with `by = join_by(Sample)` Plot over timepoints. ``` r plot<-data7%>% ggplot(aes(x=timepoint, y=protein_count, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-121-1.png) Plot as a PCA. ``` r pca_data <- data7 %>% dplyr::select(c(starts_with("Peve"), colony_id_corr, timepoint)) # Identify numeric columns numeric_cols <- sapply(pca_data, is.numeric) # Among numeric columns, find those with non-zero sum non_zero_cols <- colSums(pca_data[, numeric_cols]) != 0 # Combine non-numeric columns with numeric columns that have non-zero sum pca_data_cleaned <- cbind( pca_data[, !numeric_cols], # All non-numeric columns pca_data[, numeric_cols][, non_zero_cols] # Numeric columns with non-zero sum ) ``` ``` r scaled.pca<-prcomp(pca_data_cleaned%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_cleaned%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_cleaned, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_cleaned, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 13276 0.11776 1.5127 0.049 * ## Residual 34 99463 0.88224 ## Total 37 112739 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 View by timepoint ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_cleaned, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=FALSE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-125-1.png) Which genes are driving this? Run PLSDA and VIP. ``` r #assigning datasets X <- pca_data_cleaned levels(as.factor(X$timepoint)) ``` ## [1] "TP1" "TP2" "TP3" "TP4" ``` r Y <- as.factor(X$timepoint) #select treatment names Y ``` ## [1] TP1 TP2 TP3 TP4 TP1 TP2 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 ## [20] TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 TP1 TP2 TP3 TP4 ## Levels: TP1 TP2 TP3 TP4 ``` r X<-X%>%dplyr::select(where(is.numeric)) #pull only data columns # run PLSDA MyResult.plsda <- plsda(X,Y) # 1 Run the method plotIndiv(MyResult.plsda, ind.names = FALSE, legend=TRUE, legend.title = "Protein Catabolism", ellipse = FALSE, title="", style = "graphics", centroid=FALSE, point.lwd = 2, cex=2) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-126-1.png) Extract VIPs. ``` r #extract treatment_VIP <- PLSDA.VIP(MyResult.plsda) treatment_VIP_df <- as.data.frame(treatment_VIP[["tab"]]) treatment_VIP_df ``` ## VIP ## Peve_00018490 3.06498049 ## Peve_00018054 2.05282339 ## Peve_00037378 2.04938253 ## Peve_00027681 2.03613187 ## Peve_00024931 2.02085043 ## Peve_00036772 2.00298074 ## Peve_00018227 1.97604709 ## Peve_00004777 1.97421949 ## Peve_00024597 1.97271171 ## Peve_00020473 1.97055666 ## Peve_00011615 1.92519961 ## Peve_00031002 1.89933623 ## Peve_00037730 1.89436579 ## Peve_00022347 1.89247544 ## Peve_00025769 1.89218362 ## Peve_00006912 1.87293621 ## Peve_00042041 1.86820134 ## Peve_00015639 1.85950779 ## Peve_00007481 1.85164689 ## Peve_00042452 1.85132710 ## Peve_00001976 1.84772057 ## Peve_00021850 1.84443612 ## Peve_00021035 1.84441284 ## Peve_00029784 1.84272455 ## Peve_00036049 1.84132579 ## Peve_00024484 1.82347961 ## Peve_00009831 1.81874974 ## Peve_00008454 1.81092394 ## Peve_00016713 1.80932283 ## Peve_00001919 1.79946546 ## Peve_00014690 1.79238987 ## Peve_00041796 1.79063916 ## Peve_00033018 1.78803338 ## Peve_00017138 1.78066897 ## Peve_00009023 1.77987801 ## Peve_00027191 1.77886169 ## Peve_00038744 1.77855919 ## Peve_00028216 1.77794518 ## Peve_00027794 1.77355399 ## Peve_00013515 1.76686933 ## Peve_00015392 1.76193991 ## Peve_00034340 1.76173433 ## Peve_00014691 1.75247770 ## Peve_00018396 1.74884374 ## Peve_00025617 1.74435874 ## Peve_00005383 1.74252262 ## Peve_00044500 1.73563113 ## Peve_00028686 1.73369725 ## Peve_00026905 1.73146658 ## Peve_00013647 1.73110011 ## Peve_00030578 1.72793561 ## Peve_00008634 1.71968949 ## Peve_00027763 1.71910512 ## Peve_00001617 1.70621435 ## Peve_00025849 1.70404941 ## Peve_00040575 1.70139907 ## Peve_00040149 1.70031086 ## Peve_00002619 1.70019017 ## Peve_00036046 1.69602841 ## Peve_00004501 1.69580445 ## Peve_00012059 1.69060903 ## Peve_00034872 1.68329819 ## Peve_00004103 1.68305299 ## Peve_00007502 1.68214370 ## Peve_00023937 1.67757589 ## Peve_00030742 1.67705820 ## Peve_00042557 1.67607248 ## Peve_00001961 1.67562432 ## Peve_00045083 1.67548405 ## Peve_00038813 1.67409972 ## Peve_00028440 1.67208196 ## Peve_00043739 1.66306145 ## Peve_00002788 1.66163660 ## Peve_00018370 1.66069319 ## Peve_00001955 1.65964674 ## Peve_00035872 1.65916357 ## Peve_00024930 1.64951496 ## Peve_00031587 1.64879394 ## Peve_00019311 1.64770643 ## Peve_00001083 1.64725843 ## Peve_00027793 1.64507217 ## Peve_00013807 1.64434920 ## Peve_00001194 1.64095762 ## Peve_00036939 1.63999568 ## Peve_00032663 1.63333853 ## Peve_00038883 1.63237520 ## Peve_00031062 1.63125497 ## Peve_00018573 1.63011360 ## Peve_00032286 1.62915067 ## Peve_00007487 1.62849062 ## Peve_00002672 1.62796051 ## Peve_00040095 1.62638158 ## Peve_00019417 1.62170786 ## Peve_00000679 1.62013474 ## Peve_00005967 1.61866347 ## Peve_00038461 1.61494499 ## Peve_00028111 1.61467677 ## Peve_00037585 1.61338090 ## Peve_00008477 1.61325521 ## Peve_00032881 1.61178729 ## Peve_00038425 1.61124145 ## Peve_00001830 1.61102835 ## Peve_00026304 1.61027545 ## Peve_00039938 1.60821543 ## Peve_00028239 1.60472002 ## Peve_00028567 1.60057287 ## Peve_00036347 1.60041950 ## Peve_00037743 1.59935592 ## Peve_00026069 1.59703762 ## Peve_00015572 1.59633854 ## Peve_00011565 1.59581224 ## Peve_00036953 1.59572411 ## Peve_00026301 1.59524808 ## Peve_00025206 1.59497506 ## Peve_00000442 1.59350986 ## Peve_00024447 1.59140849 ## Peve_00025233 1.58838503 ## Peve_00001903 1.58468847 ## Peve_00014579 1.58377891 ## Peve_00020107 1.58366253 ## Peve_00036261 1.58363154 ## Peve_00040861 1.58162470 ## Peve_00034825 1.58132517 ## Peve_00003830 1.58052259 ## Peve_00015496 1.57898719 ## Peve_00021433 1.57639699 ## Peve_00015573 1.57279153 ## Peve_00030194 1.57241601 ## Peve_00004552 1.57228613 ## Peve_00041732 1.57190521 ## Peve_00045307 1.57166290 ## Peve_00004144 1.57000445 ## Peve_00017066 1.56937752 ## Peve_00008604 1.56791701 ## Peve_00022762 1.56270564 ## Peve_00014687 1.55831049 ## Peve_00017518 1.55407209 ## Peve_00035269 1.55258115 ## Peve_00006508 1.55240222 ## Peve_00042748 1.54975816 ## Peve_00034064 1.54496078 ## Peve_00038683 1.54413978 ## Peve_00005685 1.54324327 ## Peve_00007448 1.53708373 ## Peve_00044115 1.53509200 ## Peve_00008866 1.53417738 ## Peve_00038115 1.53383467 ## Peve_00011734 1.52820942 ## Peve_00034437 1.52794165 ## Peve_00007851 1.52770278 ## Peve_00006523 1.52521860 ## Peve_00030173 1.52471286 ## Peve_00025770 1.51759577 ## Peve_00005388 1.51632239 ## Peve_00028944 1.51466486 ## Peve_00011444 1.51159697 ## Peve_00020723 1.51042904 ## Peve_00004505 1.50833166 ## Peve_00018194 1.50805796 ## Peve_00038504 1.50755883 ## Peve_00012697 1.50339326 ## Peve_00040173 1.50311136 ## Peve_00036468 1.50310121 ## Peve_00028668 1.50281127 ## Peve_00004101 1.50242812 ## Peve_00004331 1.50159386 ## Peve_00033530 1.50132849 ## Peve_00010943 1.50129196 ## Peve_00041076 1.49940278 ## Peve_00029461 1.49625049 ## Peve_00010449 1.49540132 ## Peve_00024010 1.49481337 ## Peve_00015472 1.49468309 ## Peve_00024829 1.49467281 ## Peve_00000528 1.49465313 ## Peve_00006198 1.49019475 ## Peve_00037936 1.48820941 ## Peve_00035339 1.48786016 ## Peve_00002417 1.48741426 ## Peve_00029364 1.48481500 ## Peve_00013233 1.48457792 ## Peve_00031597 1.48445433 ## Peve_00032517 1.48426485 ## 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Peve_00036433 0.61837467 ## Peve_00008646 0.61807779 ## Peve_00028055 0.61750049 ## Peve_00036951 0.61699088 ## Peve_00016227 0.61698381 ## Peve_00028395 0.61576382 ## Peve_00018582 0.61551971 ## Peve_00017133 0.61438146 ## Peve_00038677 0.61437131 ## Peve_00038011 0.61399736 ## Peve_00002893 0.61379524 ## Peve_00014574 0.61331976 ## Peve_00037391 0.61281359 ## Peve_00042523 0.61180413 ## Peve_00016848 0.61172143 ## Peve_00030244 0.61011233 ## Peve_00000388 0.60988721 ## Peve_00013265 0.60938943 ## Peve_00000728 0.60888098 ## Peve_00041884 0.60809208 ## Peve_00014520 0.60790969 ## Peve_00019392 0.60788913 ## Peve_00001666 0.60773234 ## Peve_00030580 0.60756197 ## Peve_00030765 0.60746894 ## Peve_00038556 0.60737608 ## Peve_00000517 0.60736090 ## Peve_00028238 0.60735299 ## Peve_00023801 0.60720152 ## Peve_00005986 0.60675198 ## Peve_00021653 0.60631011 ## Peve_00024781 0.60607675 ## Peve_00003318 0.60572195 ## Peve_00015299 0.60553652 ## Peve_00023169 0.60504201 ## Peve_00020562 0.60430007 ## Peve_00023815 0.60299672 ## Peve_00021434 0.60211916 ## Peve_00038690 0.60193443 ## Peve_00028050 0.60045257 ## Peve_00010542 0.59993067 ## Peve_00016952 0.59964097 ## Peve_00004075 0.59835352 ## Peve_00004656 0.59790942 ## Peve_00041682 0.59662365 ## Peve_00034143 0.59510998 ## Peve_00019972 0.59488764 ## Peve_00018589 0.59428200 ## Peve_00005192 0.59396605 ## Peve_00038452 0.59307854 ## Peve_00003165 0.59254425 ## Peve_00027597 0.59227111 ## Peve_00023998 0.59163172 ## Peve_00031605 0.59002320 ## Peve_00035632 0.59001479 ## Peve_00035262 0.58990733 ## Peve_00040727 0.58923734 ## Peve_00004502 0.58901032 ## Peve_00029064 0.58899626 ## Peve_00033913 0.58817199 ## Peve_00003353 0.58788852 ## Peve_00015097 0.58784460 ## Peve_00004108 0.58764293 ## Peve_00013211 0.58751064 ## Peve_00028894 0.58650891 ## Peve_00041108 0.58591540 ## Peve_00018214 0.58510232 ## Peve_00026252 0.58502694 ## Peve_00002932 0.58470647 ## Peve_00011437 0.58446273 ## Peve_00002768 0.58407205 ## Peve_00038457 0.58406072 ## Peve_00020280 0.58388770 ## Peve_00027836 0.58339938 ## Peve_00043693 0.58247724 ## Peve_00040879 0.58187607 ## Peve_00018216 0.58182095 ## Peve_00032018 0.58163339 ## Peve_00026381 0.58162482 ## Peve_00019201 0.58110585 ## Peve_00028007 0.58109024 ## Peve_00028684 0.58088969 ## Peve_00012515 0.58082419 ## Peve_00014052 0.57978251 ## Peve_00032308 0.57971211 ## Peve_00038867 0.57930185 ## Peve_00011907 0.57673916 ## Peve_00033726 0.57632432 ## Peve_00008915 0.57604820 ## Peve_00028933 0.57569488 ## Peve_00005270 0.57568378 ## Peve_00043468 0.57450708 ## Peve_00037895 0.57345193 ## Peve_00035322 0.57290420 ## Peve_00025232 0.57224649 ## Peve_00022337 0.57190894 ## Peve_00007274 0.57139531 ## Peve_00012120 0.57138391 ## Peve_00018604 0.57125831 ## Peve_00044235 0.57060579 ## Peve_00034984 0.57022122 ## Peve_00007823 0.56829178 ## Peve_00028899 0.56714298 ## Peve_00039097 0.56700575 ## Peve_00039762 0.56683071 ## Peve_00024868 0.56666561 ## Peve_00014885 0.56645307 ## Peve_00023180 0.56585510 ## Peve_00027684 0.56485675 ## Peve_00029710 0.56435197 ## Peve_00004329 0.56431669 ## Peve_00041493 0.56362704 ## Peve_00007283 0.56292509 ## Peve_00007162 0.56280964 ## Peve_00027471 0.56248955 ## Peve_00000377 0.56177765 ## Peve_00011880 0.55996300 ## Peve_00007827 0.55905057 ## Peve_00037303 0.55782987 ## Peve_00014064 0.55773459 ## Peve_00020617 0.55631906 ## Peve_00007071 0.55565309 ## Peve_00012468 0.55555972 ## Peve_00020728 0.55511030 ## Peve_00011697 0.55456965 ## Peve_00032472 0.55454967 ## Peve_00015747 0.55422478 ## Peve_00040031 0.55355291 ## Peve_00017665 0.55348056 ## Peve_00023794 0.55340351 ## Peve_00000980 0.55258278 ## Peve_00024769 0.55202506 ## Peve_00014728 0.55174126 ## Peve_00043965 0.55125503 ## Peve_00000948 0.55105601 ## Peve_00038316 0.55084720 ## Peve_00022680 0.54993011 ## Peve_00003698 0.54920268 ## Peve_00017014 0.54919164 ## Peve_00043307 0.54810498 ## Peve_00028918 0.54809995 ## Peve_00022336 0.54809314 ## Peve_00013823 0.54728811 ## Peve_00040739 0.54660793 ## Peve_00023659 0.54634510 ## Peve_00001702 0.54566870 ## Peve_00026209 0.54548127 ## Peve_00034133 0.54424881 ## Peve_00020719 0.54284075 ## Peve_00012847 0.54273850 ## Peve_00019479 0.54215311 ## Peve_00004816 0.54205130 ## Peve_00024497 0.54127435 ## Peve_00044240 0.54095139 ## Peve_00041924 0.54094312 ## Peve_00007344 0.54055269 ## Peve_00012295 0.53924347 ## Peve_00018854 0.53849357 ## Peve_00011228 0.53696888 ## Peve_00016633 0.53636262 ## Peve_00025753 0.53587785 ## Peve_00009247 0.53366903 ## Peve_00009649 0.53332576 ## Peve_00043854 0.53280919 ## Peve_00034349 0.53258103 ## Peve_00043508 0.53243022 ## Peve_00010187 0.53209566 ## Peve_00032963 0.53064060 ## Peve_00032880 0.52968189 ## Peve_00006248 0.52926316 ## Peve_00033398 0.52907720 ## Peve_00002203 0.52864461 ## Peve_00036063 0.52861925 ## Peve_00026787 0.52825705 ## Peve_00029557 0.52701295 ## Peve_00039170 0.52605623 ## Peve_00024392 0.52600545 ## Peve_00004283 0.52502081 ## Peve_00003462 0.52465993 ## Peve_00027481 0.52453544 ## Peve_00006865 0.52445933 ## Peve_00033059 0.52409686 ## Peve_00036783 0.52389815 ## Peve_00026529 0.52333206 ## Peve_00015718 0.52204425 ## Peve_00027138 0.52076792 ## Peve_00022976 0.52075624 ## Peve_00032618 0.52070774 ## Peve_00015484 0.52068956 ## Peve_00017070 0.52008605 ## Peve_00030857 0.51990731 ## Peve_00028935 0.51932992 ## Peve_00005318 0.51923154 ## Peve_00034973 0.51896347 ## Peve_00029427 0.51800361 ## Peve_00038685 0.51796849 ## Peve_00012805 0.51757320 ## Peve_00019462 0.51752531 ## Peve_00018650 0.51614521 ## Peve_00001368 0.51596136 ## Peve_00045005 0.51549669 ## Peve_00020571 0.51525909 ## Peve_00041173 0.51525325 ## Peve_00017111 0.51399336 ## Peve_00038636 0.51310675 ## Peve_00036668 0.51231389 ## Peve_00033309 0.51184033 ## Peve_00043840 0.51109545 ## Peve_00010384 0.51049727 ## Peve_00038511 0.51032490 ## Peve_00001170 0.51006737 ## Peve_00041319 0.50935878 ## Peve_00030531 0.50930115 ## Peve_00017527 0.50725897 ## Peve_00005517 0.50495401 ## Peve_00022294 0.50398925 ## Peve_00020480 0.50266393 ## Peve_00031568 0.50168158 ## Peve_00032841 0.50128226 ## Peve_00016373 0.50100057 ## Peve_00017847 0.50050937 ## Peve_00029774 0.50035083 ## Peve_00011581 0.50022694 ## Peve_00012804 0.49733018 ## Peve_00018127 0.49707998 ## Peve_00003821 0.49593019 ## Peve_00022618 0.49478112 ## Peve_00014123 0.49459250 ## Peve_00004413 0.49429175 ## Peve_00043581 0.49373254 ## Peve_00007828 0.49325647 ## Peve_00004971 0.49310971 ## Peve_00013177 0.49309047 ## Peve_00043672 0.49274669 ## Peve_00039650 0.49266009 ## Peve_00025258 0.49249279 ## Peve_00003344 0.49234498 ## Peve_00019974 0.49225453 ## Peve_00030694 0.49216198 ## Peve_00042684 0.49110332 ## Peve_00008596 0.49093547 ## Peve_00038623 0.49025212 ## Peve_00038116 0.49017009 ## Peve_00004531 0.48940558 ## Peve_00034622 0.48897410 ## Peve_00020438 0.48896505 ## Peve_00019720 0.48872778 ## Peve_00002793 0.48767520 ## Peve_00022969 0.48761671 ## Peve_00018548 0.48742052 ## Peve_00032498 0.48680309 ## Peve_00012990 0.48379488 ## Peve_00011589 0.48267292 ## Peve_00019897 0.48233967 ## Peve_00036031 0.48124617 ## Peve_00025174 0.47894220 ## Peve_00040110 0.47710808 ## Peve_00012668 0.47611332 ## Peve_00016115 0.47601397 ## Peve_00034835 0.47569741 ## Peve_00024427 0.47532102 ## Peve_00042015 0.47494075 ## Peve_00007275 0.47385903 ## Peve_00012157 0.47348382 ## Peve_00023941 0.47335731 ## Peve_00001377 0.47279001 ## Peve_00043368 0.47274875 ## Peve_00028330 0.47174335 ## Peve_00041559 0.47148311 ## Peve_00043666 0.47132969 ## Peve_00033990 0.47077790 ## Peve_00025470 0.47075039 ## Peve_00000455 0.47054359 ## Peve_00014515 0.47012345 ## Peve_00045086 0.46962362 ## Peve_00040295 0.46962339 ## Peve_00019554 0.46844698 ## Peve_00043979 0.46833539 ## Peve_00031737 0.46827315 ## Peve_00030590 0.46798173 ## Peve_00029545 0.46795516 ## Peve_00008210 0.46782159 ## Peve_00016229 0.46771450 ## Peve_00006670 0.46770388 ## Peve_00006672 0.46764776 ## Peve_00001233 0.46659003 ## Peve_00011224 0.46573795 ## Peve_00002586 0.46255779 ## Peve_00009240 0.46251188 ## Peve_00030617 0.45954742 ## Peve_00033109 0.45843269 ## Peve_00023753 0.45600538 ## Peve_00007010 0.45589818 ## Peve_00007299 0.45501187 ## Peve_00011817 0.45387353 ## Peve_00017964 0.45363240 ## Peve_00029082 0.45311557 ## Peve_00015399 0.45274995 ## Peve_00028932 0.45266948 ## Peve_00002289 0.45124926 ## Peve_00024526 0.45098947 ## Peve_00008287 0.45004240 ## Peve_00021808 0.44982620 ## Peve_00021843 0.44982620 ## Peve_00023182 0.44982620 ## Peve_00029635 0.44827885 ## Peve_00036861 0.44775242 ## Peve_00015174 0.44642678 ## Peve_00010061 0.44577879 ## Peve_00004872 0.44510786 ## Peve_00023191 0.44485659 ## Peve_00021229 0.44481000 ## Peve_00030552 0.44469438 ## Peve_00006583 0.44340499 ## Peve_00029900 0.44259099 ## Peve_00026761 0.44164360 ## Peve_00011775 0.44122516 ## Peve_00002201 0.44085487 ## Peve_00018491 0.44072542 ## Peve_00020474 0.44066597 ## Peve_00043439 0.43990174 ## Peve_00024517 0.43644961 ## Peve_00032718 0.43624178 ## Peve_00005211 0.43589257 ## Peve_00002765 0.43504675 ## Peve_00021877 0.43453026 ## Peve_00023928 0.43297812 ## Peve_00040888 0.43182512 ## Peve_00030138 0.43149496 ## Peve_00011432 0.43109041 ## Peve_00020295 0.43000355 ## Peve_00010109 0.42952697 ## Peve_00037478 0.42608846 ## Peve_00031475 0.42604699 ## Peve_00037659 0.42471713 ## Peve_00008435 0.42354150 ## Peve_00036162 0.42354150 ## Peve_00031280 0.42343862 ## Peve_00044238 0.42328641 ## Peve_00034555 0.42177107 ## Peve_00025008 0.42166983 ## Peve_00024873 0.42158428 ## Peve_00013636 0.42133097 ## Peve_00037420 0.41968139 ## Peve_00015469 0.41883354 ## Peve_00009077 0.41878801 ## Peve_00001069 0.41820506 ## Peve_00036030 0.41764753 ## Peve_00040074 0.41742568 ## Peve_00028109 0.41732255 ## Peve_00044642 0.41512717 ## Peve_00018293 0.41476105 ## Peve_00002789 0.41399189 ## Peve_00025752 0.41397971 ## Peve_00024433 0.41329045 ## Peve_00017878 0.41216481 ## Peve_00000657 0.41127455 ## Peve_00011839 0.41014906 ## Peve_00018120 0.40983696 ## Peve_00024897 0.40983015 ## Peve_00021028 0.40921383 ## Peve_00019068 0.40903823 ## Peve_00017219 0.40840368 ## Peve_00018661 0.40781903 ## Peve_00034956 0.40675855 ## Peve_00017861 0.40490530 ## Peve_00012049 0.40377499 ## Peve_00015301 0.40061993 ## Peve_00042311 0.40020155 ## Peve_00008668 0.40014800 ## Peve_00008285 0.39945419 ## Peve_00014513 0.39679841 ## Peve_00012123 0.39509580 ## Peve_00044252 0.39362691 ## Peve_00024576 0.39342582 ## Peve_00040504 0.38920790 ## Peve_00011736 0.38857268 ## Peve_00000415 0.38681101 ## Peve_00014523 0.38590765 ## Peve_00044637 0.38511040 ## Peve_00019969 0.38334799 ## Peve_00007291 0.38179812 ## Peve_00041688 0.38142968 ## Peve_00026771 0.38018250 ## Peve_00034827 0.38009890 ## Peve_00026217 0.37631509 ## Peve_00022871 0.37255642 ## Peve_00000953 0.37125088 ## Peve_00002844 0.37111384 ## Peve_00019862 0.37098250 ## Peve_00030726 0.36903573 ## Peve_00020357 0.36893761 ## Peve_00016232 0.36832452 ## Peve_00004058 0.36777282 ## Peve_00043438 0.36615564 ## Peve_00022705 0.36518598 ## Peve_00036241 0.36426745 ## Peve_00038687 0.36418860 ## Peve_00021691 0.36149589 ## Peve_00033910 0.35923005 ## Peve_00041219 0.35808985 ## Peve_00040596 0.35599928 ## Peve_00007153 0.35477729 ## Peve_00018523 0.35444788 ## Peve_00039696 0.35299820 ## Peve_00038168 0.35234531 ## Peve_00022754 0.35122770 ## Peve_00023940 0.35120256 ## Peve_00025621 0.35095901 ## Peve_00039319 0.34966678 ## Peve_00016024 0.34716052 ## Peve_00007246 0.34616714 ## Peve_00043422 0.34582003 ## Peve_00041865 0.34537462 ## Peve_00016231 0.34471357 ## Peve_00015354 0.34410218 ## Peve_00021342 0.34280234 ## Peve_00020289 0.34269843 ## Peve_00018620 0.34227779 ## Peve_00032962 0.34204744 ## Peve_00030186 0.34111959 ## Peve_00003869 0.33746589 ## Peve_00003272 0.33709785 ## Peve_00016230 0.33651305 ## Peve_00009378 0.33528113 ## Peve_00042889 0.33405170 ## Peve_00014581 0.33163261 ## Peve_00011231 0.33067322 ## Peve_00026609 0.33059912 ## Peve_00039037 0.33050999 ## Peve_00000413 0.32972837 ## Peve_00001928 0.32899981 ## Peve_00015675 0.32843469 ## Peve_00027598 0.32789179 ## Peve_00017200 0.32784951 ## Peve_00043883 0.32475310 ## Peve_00029762 0.32297967 ## Peve_00023391 0.32202290 ## Peve_00001521 0.32044870 ## Peve_00034362 0.31991950 ## Peve_00000555 0.31858927 ## Peve_00038653 0.31713284 ## Peve_00040637 0.31712998 ## Peve_00033485 0.31702877 ## Peve_00030239 0.31671248 ## Peve_00012799 0.31280097 ## Peve_00045267 0.31198038 ## Peve_00019991 0.31015695 ## Peve_00026021 0.31013915 ## Peve_00024296 0.30916695 ## Peve_00041590 0.30675856 ## Peve_00015739 0.30561047 ## Peve_00020287 0.30494849 ## Peve_00015061 0.30085117 ## Peve_00025601 0.30013957 ## Peve_00023800 0.29736092 ## Peve_00041013 0.29450757 ## Peve_00007298 0.29446728 ## Peve_00015733 0.29424727 ## Peve_00016351 0.29156819 ## Peve_00014770 0.29116081 ## Peve_00020754 0.29000302 ## Peve_00012776 0.28866085 ## Peve_00038916 0.28618563 ## Peve_00021137 0.28369979 ## Peve_00007826 0.28362434 ## Peve_00003352 0.28322773 ## Peve_00014717 0.28074680 ## Peve_00007816 0.28063172 ## Peve_00015638 0.27973767 ## Peve_00017962 0.27759987 ## Peve_00014009 0.27732676 ## Peve_00005214 0.27684152 ## Peve_00027228 0.27442355 ## Peve_00011222 0.27441514 ## Peve_00027611 0.27294679 ## Peve_00003842 0.26902475 ## Peve_00016233 0.26798069 ## Peve_00041218 0.26636508 ## Peve_00022991 0.26556225 ## Peve_00010963 0.26471264 ## Peve_00001372 0.26197802 ## Peve_00025595 0.26177850 ## Peve_00031111 0.26077696 ## Peve_00019546 0.26035788 ## Peve_00033331 0.26011765 ## Peve_00010237 0.25905037 ## Peve_00044804 0.25533521 ## Peve_00037335 0.25451961 ## Peve_00034993 0.25358062 ## Peve_00041591 0.25349658 ## Peve_00030175 0.25167117 ## Peve_00006448 0.24432656 ## Peve_00036455 0.24213150 ## Peve_00016238 0.24073195 ## Peve_00032707 0.24069614 ## Peve_00009264 0.23984616 ## Peve_00000556 0.23720216 ## Peve_00043033 0.23330872 ## Peve_00015845 0.23309800 ## Peve_00022244 0.23232447 ## Peve_00026861 0.23182108 ## Peve_00032406 0.23113122 ## Peve_00030124 0.22628006 ## Peve_00027912 0.22444043 ## Peve_00043853 0.22353301 ## Peve_00026773 0.22301750 ## Peve_00002595 0.22176822 ## Peve_00007361 0.22164508 ## Peve_00012806 0.22127038 ## Peve_00012011 0.22096345 ## Peve_00012381 0.22041929 ## Peve_00021149 0.21864634 ## Peve_00038502 0.21671894 ## Peve_00004701 0.21142463 ## Peve_00007613 0.20855701 ## Peve_00022613 0.20680651 ## Peve_00035325 0.20525029 ## Peve_00044539 0.20396723 ## Peve_00009736 0.19897833 ## Peve_00033955 0.19099653 ## Peve_00020230 0.18934737 ## Peve_00001379 0.18705686 ## Peve_00033894 0.18656616 ## Peve_00003982 0.18570115 ## Peve_00028320 0.18541912 ## Peve_00012110 0.17442363 ## Peve_00015267 0.17434077 ## Peve_00015676 0.17137009 ## Peve_00026600 0.16992287 ## Peve_00031353 0.16933743 ## Peve_00030183 0.16850801 ## Peve_00012641 0.16554797 ## Peve_00025812 0.16458605 ## Peve_00041730 0.15715252 ## Peve_00028699 0.15668495 ## Peve_00000772 0.15406781 ## Peve_00043420 0.15278738 ## Peve_00001182 0.14394006 ## Peve_00045336 0.14357338 ## Peve_00041704 0.14000409 ## Peve_00043653 0.13933860 ## Peve_00014518 0.13797042 ## Peve_00035114 0.13659442 ## Peve_00013230 0.13283718 ## Peve_00028110 0.13172107 ## Peve_00036351 0.12630078 ## Peve_00027038 0.12214524 ## Peve_00037077 0.12118753 ## Peve_00019968 0.11745724 ## Peve_00027685 0.11557751 ## Peve_00040729 0.11550794 ## Peve_00013229 0.10515108 ## Peve_00025727 0.10194725 ## Peve_00008288 0.08825430 ## Peve_00027035 0.08410330 ## Peve_00005459 0.06988039 ## Peve_00012142 0.06848180 ## Peve_00041586 0.05662385 ## Peve_00013628 0.02953256 ``` r # Converting row names to column treatment_VIP_table <- rownames_to_column(treatment_VIP_df, var = "Gene") #filter for VIP > 1 treatment_VIP_1 <- treatment_VIP_table %>% filter(VIP >= 1.5) #plot VIP_list_plot<-treatment_VIP_1 %>% arrange(VIP) %>% ggplot( aes(x = VIP, y = reorder(Gene,VIP,sum))) + geom_point() + ylab("Gene") + xlab("VIP Score") + ggtitle("Protein Catabolism") + theme_bw() + theme(panel.border = element_rect(linetype = "solid", color = "black"), panel.grid.major = element_blank(), #Makes background theme white panel.grid.minor = element_blank(), axis.line = element_line(colour = "black"));VIP_list_plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-127-1.png) ``` r treatment_VIP_1 %>% arrange(VIP) ``` ## Gene VIP ## 1 Peve_00010943 1.501292 ## 2 Peve_00033530 1.501328 ## 3 Peve_00004331 1.501594 ## 4 Peve_00004101 1.502428 ## 5 Peve_00028668 1.502811 ## 6 Peve_00036468 1.503101 ## 7 Peve_00040173 1.503111 ## 8 Peve_00012697 1.503393 ## 9 Peve_00038504 1.507559 ## 10 Peve_00018194 1.508058 ## 11 Peve_00004505 1.508332 ## 12 Peve_00020723 1.510429 ## 13 Peve_00011444 1.511597 ## 14 Peve_00028944 1.514665 ## 15 Peve_00005388 1.516322 ## 16 Peve_00025770 1.517596 ## 17 Peve_00030173 1.524713 ## 18 Peve_00006523 1.525219 ## 19 Peve_00007851 1.527703 ## 20 Peve_00034437 1.527942 ## 21 Peve_00011734 1.528209 ## 22 Peve_00038115 1.533835 ## 23 Peve_00008866 1.534177 ## 24 Peve_00044115 1.535092 ## 25 Peve_00007448 1.537084 ## 26 Peve_00005685 1.543243 ## 27 Peve_00038683 1.544140 ## 28 Peve_00034064 1.544961 ## 29 Peve_00042748 1.549758 ## 30 Peve_00006508 1.552402 ## 31 Peve_00035269 1.552581 ## 32 Peve_00017518 1.554072 ## 33 Peve_00014687 1.558310 ## 34 Peve_00022762 1.562706 ## 35 Peve_00008604 1.567917 ## 36 Peve_00017066 1.569378 ## 37 Peve_00004144 1.570004 ## 38 Peve_00045307 1.571663 ## 39 Peve_00041732 1.571905 ## 40 Peve_00004552 1.572286 ## 41 Peve_00030194 1.572416 ## 42 Peve_00015573 1.572792 ## 43 Peve_00021433 1.576397 ## 44 Peve_00015496 1.578987 ## 45 Peve_00003830 1.580523 ## 46 Peve_00034825 1.581325 ## 47 Peve_00040861 1.581625 ## 48 Peve_00036261 1.583632 ## 49 Peve_00020107 1.583663 ## 50 Peve_00014579 1.583779 ## 51 Peve_00001903 1.584688 ## 52 Peve_00025233 1.588385 ## 53 Peve_00024447 1.591408 ## 54 Peve_00000442 1.593510 ## 55 Peve_00025206 1.594975 ## 56 Peve_00026301 1.595248 ## 57 Peve_00036953 1.595724 ## 58 Peve_00011565 1.595812 ## 59 Peve_00015572 1.596339 ## 60 Peve_00026069 1.597038 ## 61 Peve_00037743 1.599356 ## 62 Peve_00036347 1.600420 ## 63 Peve_00028567 1.600573 ## 64 Peve_00028239 1.604720 ## 65 Peve_00039938 1.608215 ## 66 Peve_00026304 1.610275 ## 67 Peve_00001830 1.611028 ## 68 Peve_00038425 1.611241 ## 69 Peve_00032881 1.611787 ## 70 Peve_00008477 1.613255 ## 71 Peve_00037585 1.613381 ## 72 Peve_00028111 1.614677 ## 73 Peve_00038461 1.614945 ## 74 Peve_00005967 1.618663 ## 75 Peve_00000679 1.620135 ## 76 Peve_00019417 1.621708 ## 77 Peve_00040095 1.626382 ## 78 Peve_00002672 1.627961 ## 79 Peve_00007487 1.628491 ## 80 Peve_00032286 1.629151 ## 81 Peve_00018573 1.630114 ## 82 Peve_00031062 1.631255 ## 83 Peve_00038883 1.632375 ## 84 Peve_00032663 1.633339 ## 85 Peve_00036939 1.639996 ## 86 Peve_00001194 1.640958 ## 87 Peve_00013807 1.644349 ## 88 Peve_00027793 1.645072 ## 89 Peve_00001083 1.647258 ## 90 Peve_00019311 1.647706 ## 91 Peve_00031587 1.648794 ## 92 Peve_00024930 1.649515 ## 93 Peve_00035872 1.659164 ## 94 Peve_00001955 1.659647 ## 95 Peve_00018370 1.660693 ## 96 Peve_00002788 1.661637 ## 97 Peve_00043739 1.663061 ## 98 Peve_00028440 1.672082 ## 99 Peve_00038813 1.674100 ## 100 Peve_00045083 1.675484 ## 101 Peve_00001961 1.675624 ## 102 Peve_00042557 1.676072 ## 103 Peve_00030742 1.677058 ## 104 Peve_00023937 1.677576 ## 105 Peve_00007502 1.682144 ## 106 Peve_00004103 1.683053 ## 107 Peve_00034872 1.683298 ## 108 Peve_00012059 1.690609 ## 109 Peve_00004501 1.695804 ## 110 Peve_00036046 1.696028 ## 111 Peve_00002619 1.700190 ## 112 Peve_00040149 1.700311 ## 113 Peve_00040575 1.701399 ## 114 Peve_00025849 1.704049 ## 115 Peve_00001617 1.706214 ## 116 Peve_00027763 1.719105 ## 117 Peve_00008634 1.719689 ## 118 Peve_00030578 1.727936 ## 119 Peve_00013647 1.731100 ## 120 Peve_00026905 1.731467 ## 121 Peve_00028686 1.733697 ## 122 Peve_00044500 1.735631 ## 123 Peve_00005383 1.742523 ## 124 Peve_00025617 1.744359 ## 125 Peve_00018396 1.748844 ## 126 Peve_00014691 1.752478 ## 127 Peve_00034340 1.761734 ## 128 Peve_00015392 1.761940 ## 129 Peve_00013515 1.766869 ## 130 Peve_00027794 1.773554 ## 131 Peve_00028216 1.777945 ## 132 Peve_00038744 1.778559 ## 133 Peve_00027191 1.778862 ## 134 Peve_00009023 1.779878 ## 135 Peve_00017138 1.780669 ## 136 Peve_00033018 1.788033 ## 137 Peve_00041796 1.790639 ## 138 Peve_00014690 1.792390 ## 139 Peve_00001919 1.799465 ## 140 Peve_00016713 1.809323 ## 141 Peve_00008454 1.810924 ## 142 Peve_00009831 1.818750 ## 143 Peve_00024484 1.823480 ## 144 Peve_00036049 1.841326 ## 145 Peve_00029784 1.842725 ## 146 Peve_00021035 1.844413 ## 147 Peve_00021850 1.844436 ## 148 Peve_00001976 1.847721 ## 149 Peve_00042452 1.851327 ## 150 Peve_00007481 1.851647 ## 151 Peve_00015639 1.859508 ## 152 Peve_00042041 1.868201 ## 153 Peve_00006912 1.872936 ## 154 Peve_00025769 1.892184 ## 155 Peve_00022347 1.892475 ## 156 Peve_00037730 1.894366 ## 157 Peve_00031002 1.899336 ## 158 Peve_00011615 1.925200 ## 159 Peve_00020473 1.970557 ## 160 Peve_00024597 1.972712 ## 161 Peve_00004777 1.974219 ## 162 Peve_00018227 1.976047 ## 163 Peve_00036772 2.002981 ## 164 Peve_00024931 2.020850 ## 165 Peve_00027681 2.036132 ## 166 Peve_00037378 2.049383 ## 167 Peve_00018054 2.052823 ## 168 Peve_00018490 3.064980 Plot important genes. ``` r plot<-data7%>% ggplot(aes(x=timepoint, y=Peve_00018490, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-128-1.png) Plot second most important ``` r plot<-data7%>% ggplot(aes(x=timepoint, y=Peve_00018054, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-129-1.png) Plot third most important ``` r plot<-data7%>% ggplot(aes(x=timepoint, y=Peve_00037378, group=colony_id_corr))+ facet_wrap(~species)+ geom_point()+ geom_line()+ theme_classic();plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-130-1.png) Look at a PCA of the differentiating genes. ``` r #extract list of VIPs vip_genes<-treatment_VIP_1%>%pull(Gene) #turn to wide format with pca_data_vips<-pca_data_cleaned%>%dplyr::select(all_of(c("timepoint", "colony_id_corr", vip_genes))) ``` ``` r scaled.pca<-prcomp(pca_data_vips%>%dplyr::select(where(is.numeric)), scale=TRUE, center=TRUE) ``` Prepare a PCA plot ``` r # scale data vegan <- scale(pca_data_vips%>%dplyr::select(where(is.numeric))) # PerMANOVA permanova<-adonis2(vegan ~ timepoint, data = pca_data_vips, method='eu') permanova ``` ## Permutation test for adonis under reduced model ## Terms added sequentially (first to last) ## Permutation: free ## Number of permutations: 999 ## ## adonis2(formula = vegan ~ timepoint, data = pca_data_vips, method = "eu") ## Df SumOfSqs R2 F Pr(>F) ## timepoint 3 1441.5 0.23191 3.4218 0.001 *** ## Residual 34 4774.5 0.76809 ## Total 37 6216.0 1.00000 ## --- ## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1 Significant differences in fatty acid beta oxidation gene expression profile between time points. View by species ``` r plot<-ggplot2::autoplot(scaled.pca, data=pca_data_vips, loadings=FALSE, colour="timepoint", loadings.label.colour="black", loadings.colour="black", loadings.label=FALSE, frame=TRUE, loadings.label.size=5, loadings.label.vjust=-1, size=5) + theme_classic()+ ggtitle("Protein Catabolism")+ theme(legend.text = element_text(size=18), legend.position="right", plot.background = element_blank(), legend.title = element_text(size=18, face="bold"), axis.text = element_text(size=18), axis.title = element_text(size=18, face="bold"));plot ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-134-1.png) Pull out PC1 score for each sample for GO term. ``` r scores1 <- scaled.pca$x scores1<-as.data.frame(scores1) scores1<-scores1%>%dplyr::select(PC1) scores1$sample<-pca_data_vips$colony_id_corr scores1$timepoint<-pca_data_vips$timepoint scores1<-scores1%>% rename(protein=PC1) scores<-left_join(scores, scores1) ``` ## Joining with `by = join_by(sample, timepoint)` ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis lipolysis fa_beta starve ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 -2.7317639 -17.737096 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 -15.4501240 -4.815820 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 0.2483462 4.222578 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 7.3058463 6.012566 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 -12.1525556 -3.705685 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 2.0021942 7.931360 ## lipids protein ## 1 -16.022543 -19.254671 ## 2 -3.371010 -3.279044 ## 3 4.226094 5.002136 ## 4 5.368042 5.989240 ## 5 -3.538633 -3.198097 ## 6 7.235845 8.335553 # Output PC1 scores for each GO term for each sample ``` r head(scores) ``` ## glycolysis sample timepoint gluconeogenesis lipolysis fa_beta starve ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 -2.7317639 -17.737096 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 -15.4501240 -4.815820 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 0.2483462 4.222578 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 7.3058463 6.012566 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 -12.1525556 -3.705685 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 2.0021942 7.931360 ## lipids protein ## 1 -16.022543 -19.254671 ## 2 -3.371010 -3.279044 ## 3 4.226094 5.002136 ## 4 5.368042 5.989240 ## 5 -3.538633 -3.198097 ## 6 7.235845 8.335553 ``` r scores<-scores%>% rename(colony_id_corr=sample) ``` # Correlate PC scores to physiology metrics ``` r #merge scores into physiology data frame head(phys) ``` ## colony_id_corr timepoint species site Host_AFDW.mg.cm2 Sym_AFDW.mg.cm2 ## 1 POR-216 TP1 Porites Mahana 4.438584 1.570576 ## 2 POR-216 TP2 Porites Mahana 8.909985 4.037532 ## 3 POR-216 TP3 Porites Mahana 8.563552 4.498444 ## 4 POR-216 TP4 Porites Mahana 3.085705 1.753038 ## 5 POR-236 TP1 Porites Mahana 3.710084 2.135899 ## 6 POR-236 TP2 Porites Mahana 7.197873 3.454979 ## Am AQY Rd Ik Ic calc.umol.cm2.hr ## 1 1.533779 0.005015782 0.7043682 305.7906 158.0865 0.002833631 ## 2 3.490678 0.008258861 1.0871774 422.6585 138.5278 1.067968298 ## 3 2.013604 0.004443360 0.8679998 453.1716 216.4949 1.020121080 ## 4 1.299239 0.003778211 0.5186043 343.8767 149.7052 0.008450092 ## 5 1.921312 0.005106034 1.0078725 376.2826 231.8497 0.038408285 ## 6 2.132806 0.003206807 0.7228886 665.0872 239.6058 1.278575133 ## cells.mgAFDW prot_mg.mgafdw Ratio_AFDW.mg.cm2 Total_Chl Total_Chl_cell ## 1 321153.85 0.3422878 0.2613636 1.2313622 3.834182e-06 ## 2 227505.83 0.3488877 0.3118383 1.1295221 4.964805e-06 ## 3 88621.15 0.3184877 0.3443918 0.9918775 1.119233e-05 ## 4 94202.90 0.3914365 0.3622921 1.0533092 1.118128e-05 ## 5 299273.10 0.4552645 0.3653618 3.1498702 1.052507e-05 ## 6 246956.52 0.3658136 0.3243243 1.2541449 5.078404e-06 ## cre.umol.mgafdw Sample ## 1 0.08464618 POR-216_TP1 ## 2 0.21351684 POR-216_TP2 ## 3 0.23139768 POR-216_TP3 ## 4 0.20885178 POR-216_TP4 ## 5 0.22508911 POR-236_TP1 ## 6 0.27360399 POR-236_TP2 ``` r head(scores) ``` ## glycolysis colony_id_corr timepoint gluconeogenesis lipolysis fa_beta ## 1 -3.423127 POR-216 TP1 -6.749172 10.646590 -2.7317639 ## 2 -13.074547 POR-216 TP2 -10.860762 6.295957 -15.4501240 ## 3 1.186372 POR-216 TP3 1.457899 -2.164272 0.2483462 ## 4 4.455591 POR-216 TP4 7.899062 -4.821601 7.3058463 ## 5 -9.380917 POR-236 TP1 -11.102232 5.090455 -12.1525556 ## 6 1.519258 POR-236 TP2 3.529447 -3.952191 2.0021942 ## starve lipids protein ## 1 -17.737096 -16.022543 -19.254671 ## 2 -4.815820 -3.371010 -3.279044 ## 3 4.222578 4.226094 5.002136 ## 4 6.012566 5.368042 5.989240 ## 5 -3.705685 -3.538633 -3.198097 ## 6 7.931360 7.235845 8.335553 ``` r main<-left_join(phys, scores) ``` ## Joining with `by = join_by(colony_id_corr, timepoint)` ``` r head(main) ``` ## colony_id_corr timepoint species site Host_AFDW.mg.cm2 Sym_AFDW.mg.cm2 ## 1 POR-216 TP1 Porites Mahana 4.438584 1.570576 ## 2 POR-216 TP2 Porites Mahana 8.909985 4.037532 ## 3 POR-216 TP3 Porites Mahana 8.563552 4.498444 ## 4 POR-216 TP4 Porites Mahana 3.085705 1.753038 ## 5 POR-236 TP1 Porites Mahana 3.710084 2.135899 ## 6 POR-236 TP2 Porites Mahana 7.197873 3.454979 ## Am AQY Rd Ik Ic calc.umol.cm2.hr ## 1 1.533779 0.005015782 0.7043682 305.7906 158.0865 0.002833631 ## 2 3.490678 0.008258861 1.0871774 422.6585 138.5278 1.067968298 ## 3 2.013604 0.004443360 0.8679998 453.1716 216.4949 1.020121080 ## 4 1.299239 0.003778211 0.5186043 343.8767 149.7052 0.008450092 ## 5 1.921312 0.005106034 1.0078725 376.2826 231.8497 0.038408285 ## 6 2.132806 0.003206807 0.7228886 665.0872 239.6058 1.278575133 ## cells.mgAFDW prot_mg.mgafdw Ratio_AFDW.mg.cm2 Total_Chl Total_Chl_cell ## 1 321153.85 0.3422878 0.2613636 1.2313622 3.834182e-06 ## 2 227505.83 0.3488877 0.3118383 1.1295221 4.964805e-06 ## 3 88621.15 0.3184877 0.3443918 0.9918775 1.119233e-05 ## 4 94202.90 0.3914365 0.3622921 1.0533092 1.118128e-05 ## 5 299273.10 0.4552645 0.3653618 3.1498702 1.052507e-05 ## 6 246956.52 0.3658136 0.3243243 1.2541449 5.078404e-06 ## cre.umol.mgafdw Sample glycolysis gluconeogenesis lipolysis fa_beta ## 1 0.08464618 POR-216_TP1 -3.423127 -6.749172 10.646590 -2.7317639 ## 2 0.21351684 POR-216_TP2 -13.074547 -10.860762 6.295957 -15.4501240 ## 3 0.23139768 POR-216_TP3 1.186372 1.457899 -2.164272 0.2483462 ## 4 0.20885178 POR-216_TP4 4.455591 7.899062 -4.821601 7.3058463 ## 5 0.22508911 POR-236_TP1 -9.380917 -11.102232 5.090455 -12.1525556 ## 6 0.27360399 POR-236_TP2 1.519258 3.529447 -3.952191 2.0021942 ## starve lipids protein ## 1 -17.737096 -16.022543 -19.254671 ## 2 -4.815820 -3.371010 -3.279044 ## 3 4.222578 4.226094 5.002136 ## 4 6.012566 5.368042 5.989240 ## 5 -3.705685 -3.538633 -3.198097 ## 6 7.931360 7.235845 8.335553 ``` r main<-main%>% dplyr::select(where(is.numeric)) head(main) ``` ## Host_AFDW.mg.cm2 Sym_AFDW.mg.cm2 Am AQY Rd Ik ## 1 4.438584 1.570576 1.533779 0.005015782 0.7043682 305.7906 ## 2 8.909985 4.037532 3.490678 0.008258861 1.0871774 422.6585 ## 3 8.563552 4.498444 2.013604 0.004443360 0.8679998 453.1716 ## 4 3.085705 1.753038 1.299239 0.003778211 0.5186043 343.8767 ## 5 3.710084 2.135899 1.921312 0.005106034 1.0078725 376.2826 ## 6 7.197873 3.454979 2.132806 0.003206807 0.7228886 665.0872 ## Ic calc.umol.cm2.hr cells.mgAFDW prot_mg.mgafdw Ratio_AFDW.mg.cm2 ## 1 158.0865 0.002833631 321153.85 0.3422878 0.2613636 ## 2 138.5278 1.067968298 227505.83 0.3488877 0.3118383 ## 3 216.4949 1.020121080 88621.15 0.3184877 0.3443918 ## 4 149.7052 0.008450092 94202.90 0.3914365 0.3622921 ## 5 231.8497 0.038408285 299273.10 0.4552645 0.3653618 ## 6 239.6058 1.278575133 246956.52 0.3658136 0.3243243 ## Total_Chl Total_Chl_cell cre.umol.mgafdw glycolysis gluconeogenesis lipolysis ## 1 1.2313622 3.834182e-06 0.08464618 -3.423127 -6.749172 10.646590 ## 2 1.1295221 4.964805e-06 0.21351684 -13.074547 -10.860762 6.295957 ## 3 0.9918775 1.119233e-05 0.23139768 1.186372 1.457899 -2.164272 ## 4 1.0533092 1.118128e-05 0.20885178 4.455591 7.899062 -4.821601 ## 5 3.1498702 1.052507e-05 0.22508911 -9.380917 -11.102232 5.090455 ## 6 1.2541449 5.078404e-06 0.27360399 1.519258 3.529447 -3.952191 ## fa_beta starve lipids protein ## 1 -2.7317639 -17.737096 -16.022543 -19.254671 ## 2 -15.4501240 -4.815820 -3.371010 -3.279044 ## 3 0.2483462 4.222578 4.226094 5.002136 ## 4 7.3058463 6.012566 5.368042 5.989240 ## 5 -12.1525556 -3.705685 -3.538633 -3.198097 ## 6 2.0021942 7.931360 7.235845 8.335553 Compute correlations ``` r # Compute correlations with rcorr corr_result <- rcorr(as.matrix(main), type = "pearson") # Extract correlation matrix and p-values cor_matrix <- corr_result$r p_matrix <- corr_result$P # Get correlation between sets of interest only terms<-c("glycolysis", "gluconeogenesis", "lipolysis", "fa_beta", "starve", "protein", "lipids") phys_var<-c("Host_AFDW.mg.cm2", "Sym_AFDW.mg.cm2", "Am", "AQY", "Rd", "Ik", "Ic", "calc.umol.cm2.hr", "cells.mgAFDW", "prot_mg.mgafdw", "Ratio_AFDW.mg.cm2", "Total_Chl", "Total_Chl_cell", "cre.umol.mgafdw") # Subset the correlation matrix subset_cor <- cor_matrix[terms, phys_var, drop = FALSE] # Subset the p-value matrix subset_p <- p_matrix[terms, phys_var, drop = FALSE] ``` Plot heatmap ``` r # Create matrix of formatted p-values p_labels <- matrix(sprintf("%.3f", subset_p), nrow = nrow(subset_p)) # Plot with corrplot corrplot(subset_cor, method = "color", # Fill squares with color col = colorRampPalette(c("blue", "white", "red"))(200), type = "full", tl.col = "black", # Text label color tl.cex = 1.1, # Label size addCoef.col = "black", # Show correlation values (optional) number.cex = 0.8, # Text size for numbers p.mat = subset_p, # p-value matrix insig = "blank", # Only show significant diag = FALSE # Hide diagonal ) ``` ![](07-Peve-energetic-state_files/figure-gfm/unnamed-chunk-139-1.png) ``` r # Overlay p-values as text manually (optional if not showing coef) #text(x = rep(1:ncol(subset_p), each = nrow(subset_p)), # y = rep(nrow(subset_p):1, ncol(subset_p)), # labels = p_labels, # cex = 0.8) ```