---
title: "kallisto"
output: html_document
date: "2023-11-27"
---

```{r setup, include=FALSE}
knitr::opts_chunk$set(echo = TRUE)
```

kallisto info and location on Raven

```{bash}

/home/shared/kallisto/kallisto
```

## Retrieving the [reference genome](https://www.ncbi.nlm.nih.gov/datasets/genome/GCF_026571515.1/)

```{bash}

curl -OJX GET "https://api.ncbi.nlm.nih.gov/datasets/v2alpha/genome/accession/GCF_026571515.1/download?include_annotation_type=GENOME_FASTA,GENOME_GFF,RNA_FASTA,CDS_FASTA,PROT_FASTA,SEQUENCE_REPORT&filename=GCF_026571515.1.zip" -H "Accept: application/zip" 
```

**need to unzip in command line and follow prompts.**

once done, move reference file folder to data folder (saves it initially to current directory: code)

```{bash}

mv ncbi_dataset/ ../data/


```

## Building Index

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto index -t 20 -i transcripts.idx ../data/ncbi_dataset/data/GCF_026571515.1/rna.fna 

```

# Quantification

### code breakdown :

`/home/shared/kallisto/kallisto` kallisto path

`quant` tells kallisto to run the quantification algorithm

`-i transcripts.idx` specifies the `-i` index file (created in previous step)

`-o ../output/kallisto/sampleID/` specifies the `-o` output directory

`-b 100` creates 100 `-b` bootstraps. Included in [Kallisto tutorial](https://pachterlab.github.io/kallisto/starting) does not influence quantification output.

`…/data/raw/sampleID_R1_001.fastq.gz` read 1 of the sample

`…/data/raw/sampleID_R2_001.fastq.gz` read 2 of the sample

## Control Sample Quantification

chunk run time: 9:16am start, 9:25am finish

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-216/ -b 100  ../data/raw/M-C-216_R1_001.fastq.gz ../data/raw/M-C-216_R2_001.fastq.gz

```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-218/ -b 100  ../data/raw/M-C-218_R1_001.fastq.gz ../data/raw/M-C-218_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-226/ -b 100  ../data/raw/M-C-226_R1_001.fastq.gz ../data/raw/M-C-226_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-306/ -b 100  ../data/raw/M-C-306_R1_001.fastq.gz ../data/raw/M-C-306_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-329/ -b 100  ../data/raw/M-C-329_R1_001.fastq.gz ../data/raw/M-C-329_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-334/ -b 100  ../data/raw/M-C-334_R1_001.fastq.gz ../data/raw/M-C-334_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-337/ -b 100  ../data/raw/M-C-337_R1_001.fastq.gz ../data/raw/M-C-337_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-339/ -b 100  ../data/raw/M-C-339_R1_001.fastq.gz ../data/raw/M-C-339_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-358/ -b 100  ../data/raw/M-C-358_R1_001.fastq.gz ../data/raw/M-C-358_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-360/ -b 100  ../data/raw/M-C-360_R1_001.fastq.gz ../data/raw/M-C-360_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-363/ -b 100  ../data/raw/M-C-363_R1_001.fastq.gz ../data/raw/M-C-363_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-373/ -b 100  ../data/raw/M-C-373_R1_001.fastq.gz ../data/raw/M-C-373_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-482/ -b 100  ../data/raw/M-C-482_R1_001.fastq.gz ../data/raw/M-C-482_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-488/ -b 100  ../data/raw/M-C-488_R1_001.fastq.gz ../data/raw/M-C-488_R2_001.fastq.gz
```

```{bash}
/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-C-193/ -b 100  ../data/raw/M-C-193_R1_001.fastq.gz ../data/raw/M-C-193_R2_001.fastq.gz
```

## Treatment Sample Quantification

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-18/ -b 100  ../data/raw/M-T-18_R1_001.fastq.gz ../data/raw/M-T-18_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-20/ -b 100  ../data/raw/M-T-20_R1_001.fastq.gz ../data/raw/M-T-20_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-22/ -b 100  ../data/raw/M-T-22_R1_001.fastq.gz ../data/raw/M-T-22_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-235/ -b 100  ../data/raw/M-T-235_R1_001.fastq.gz ../data/raw/M-T-235_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-245/ -b 100  ../data/raw/M-T-245_R1_001.fastq.gz ../data/raw/M-T-245_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-29/ -b 100  ../data/raw/M-T-29_R1_001.fastq.gz ../data/raw/M-T-29_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-31/ -b 100  ../data/raw/M-T-31_R1_001.fastq.gz ../data/raw/M-T-31_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-32/ -b 100  ../data/raw/M-T-32_R1_001.fastq.gz ../data/raw/M-T-32_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-399/ -b 100  ../data/raw/M-T-399_R1_001.fastq.gz ../data/raw/M-T-399_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-43/ -b 100  ../data/raw/M-T-43_R1_001.fastq.gz ../data/raw/M-T-43_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-44/ -b 100  ../data/raw/M-T-44_R1_001.fastq.gz ../data/raw/M-T-44_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-500/ -b 100  ../data/raw/M-T-500_R1_001.fastq.gz ../data/raw/M-T-500_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-7/ -b 100  ../data/raw/M-T-7_R1_001.fastq.gz ../data/raw/M-T-7_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-83/ -b 100  ../data/raw/M-T-83_R1_001.fastq.gz ../data/raw/M-T-83_R2_001.fastq.gz
```

```{bash}

/home/shared/kallisto_linux-v0.50.1/kallisto quant -t 10 -i transcripts.idx -o ../output/kallisto/M-T-8/ -b 100  ../data/raw/M-T-8_R1_001.fastq.gz ../data/raw/M-T-8_R2_001.fastq.gz
```

# Trinity Matrix with Kallisto Output

run time: \~1 min

```{bash}

/home/shared/trinityrnaseq-v2.12.0/util/abundance_estimates_to_matrix.pl --est_method kallisto --gene_trans_map 'none' --out_prefix kallisto --name_sample_by_basedir ../output/kallisto/**/*.tsv


```

moving results to proper directory

```{bash}

mv kallisto.isoform* ../output/
```