RNA-seq Automation

This work performs a Fastqc, generates a Genome index, genome aligning using STAR, and count matrix using subread

Start by cloning the source code:

git clone https://github.com/Ahmed-Ghobashi/RNA-seq.git
cd RNA-seq

if you are working on HPC, you could load the dependencies using the module load function

or You can create the virtual sofware environment, and download all software and dependencies. Create the new environment and specify the software to include in it.

conda create -n < name of your environment > -y -c conda-forge -c bioconda \
pandas fastqc star samtools subread

Now you can activate your environment conda activate <name of your environment>.

RNA-seq automation

In the terminal

pyhton RNA_seq.py 
-n, --name---> name of your job,default='RNA-seq_run'
-j, --job'-->default='ALL', choices=['ALL','Fastqc','Genomic_index','Count']
-sd, --save_dir--->default='results', name of the result directory
-th, --thread_core-->default='2', number of the cores
-i, --genome_index--> ask if you have genome index, default='TRUE', choices=['TRUE','FALSE'])
-gd', --genome_dir--> directory for the genome index
-fa, --fasta_dir-->directory of fasta file
-gtf, --gtf_dir-->directory of GTF file
-c, --condition-->write the sample name and replicate
-r1, --read1-->read number 1 of your sample
-p, --paired--> the sequencing is paired or single end ,default='NO',choices=['YES','NO']
-r2, --read2-->read number 2 of your sample (for paired-end only)
-s, --strandness--> 0 for unstranded reads, 1 for stranded reads and 2 for reversely stranded reads. This depends on the library used in the sequencing protocol. 
                     <Most commercial kits use standed protocol',default='1',choices=['0','1','2']

For example

 python RNA_seq.py -th 6 -gd /N/slate/aghobash/STAR/Genecod -fa /N/slate/aghobash/STAR/GRCh38.primary_assembly.genome.fa\
 -gtf /N/slate/aghobash/STAR/gencode.v29.annotation.gtf -c EV_1 -r1 GSF3258-mRNAseq-9_S19_R1_001.fastq.gz\
 -p YES -r2 GSF3258-mRNAseq-9_S19_R2_001.fastq.gz

This work depends on

FastQC - Read quality control.
STAR - Read aligner.
Samtools - SAM/BAM manipulation.
Subread - Read annotation and counting.