---
title: "05-Apul lncRNA"
author: Steven Roberts
date: "`r format(Sys.time(), '%d %B, %Y')`"  
output: 
  html_document:
    theme: readable
    highlight: zenburn
    toc: true
    toc_float: true
    number_sections: true
    code_folding: show
    code_download: true
---

```{r setup, include=FALSE}
library(knitr)
library(tidyverse)
knitr::opts_chunk$set(
  echo = TRUE,         # Display code chunks
  eval = FALSE,         # Evaluate code chunks
  warning = FALSE,     # Hide warnings
  message = FALSE,     # Hide messages
  fig.width = 6,       # Set plot width in inches
  fig.height = 4,      # Set plot height in inches
  fig.align = "center" # Align plots to the center
)
```


I already have bed and fasta of lncRNA from deep-dive expression..

```{bash}
head /home/shared/8TB_HDD_03/sr320/github/deep-dive-expression/D-Apul/output/10.1-Apul-lncRNA/*lncRNA.*
```
4. Quantify lncRNA expression

Use featureCounts from the Subread package to count the reads mapped to your lncRNAs. Since you already have a BED file, convert it to a GTF file or use tools that accept BED directly.

Convert BED to GTF (if required)

Use bedToGtf.py or similar scripts to convert the BED file to a GTF file.

```{bash}
awk 'BEGIN{OFS="\t"; count=1} {printf "%s\t.\tlncRNA\t%d\t%d\t.\t+\t.\tgene_id \"lncRNA_%03d\";\n", $1, $2, $3, count++;}' /home/shared/8TB_HDD_03/sr320/github/deep-dive-expression/D-Apul/output/10.1-Apul-lncRNA/Apul_lncRNA.bed \
> ../output/05-Apul-lncRNA/lncRNAs.gtf
```

```{bash}
head ../data/Apulchra-genome.gff
```



```{r, engine='bash'}
head ../output/05-Apul-lncRNA/lncRNAs.gtf
```
```{bash}
awk '{if(NF<9) print "Error in line:", NR, $0}' ../output/05-Apul-lncRNA/lncRNAs.gtf
```


```{bash}
head -n 1 ../output/05-Apul-lncRNA/lncRNAs.gtf > ../output/05-Apul-lncRNA/test_lncRNAs.gtf
```

```{bash}
head ../output/05-Apul-lncRNA/test_lncRNAs.gtf
```



```{r, engine='bash'}
/home/shared/subread-2.0.5-Linux-x86_64/bin/featureCounts \
-T 42 \
-a ../output/05-Apul-lncRNA/lncRNAs.gtf \
-o ../output/05-Apul-lncRNA/counts.txt \
-t lncRNA \
-g gene_id \
-p \
../data/*sorted.bam
```
```{bash}
samtools view -H ../data/2C1.sorted.bam | grep '@SQ'
```


```{r, engine='bash'}
find ../data/*sorted.bam \
| xargs basename -s .sorted.bam | xargs -I{} \
/home/shared/stringtie-2.2.1.Linux_x86_64/stringtie \
-p 42 \
-eB \
-G ../output/05-Apul-lncRNA/lncRNAs.gtf \
-o ../output/05-Apul-lncRNA/{}.gtf \
../data/{}.sorted.bam
```

