RNA-Seq Flow is the script written in python snakemake format which starts from the raw fastq files and ends all the way to give you gene and Isoform level count using RSEM, to Increase the mapping effeciency, it does 2nd pass STAR Allignment by indexing the genome again using the merged SJ.0UT.tab files from the 1st pass. for the quality control it does Fastqc as well as does trimming though Trim-Galore.

• FastQC (A quality control tool for high throughput sequence data)

• Trim-galore (Automates quality control and adapter trimming of fastq files)

• STAR (Spliced aware ultrafast transcript alligner to refernece genome)

• Picard (Cammand line set tool to manipulate high-throughput sequencing data)

• RNA-SeQC (Qualiy Control metrices of RNA-Seq data)

• RSEM (Accurate quantification of gene and isoform expression levels from RNA-Seq data)

• Install anaconda or load it if it's already on your server

• conda create --name rnaseq-env

• source activate rnaseq-env

• conda install -c bioconda star

• conda install -c bioconda fastqc

• conda install -c bioconda rsem

 STAR  --runMode genomeGenerate --runThreadN 24 --genomeDir ./ --genomeFastaFiles hg38.fa   --sjdbGTFfile gencode.v30.annotation.gtf 

 python3 fastqc-summary -s $INDIR > "QC_Report.txt"

python3 collapse_annotation.py gencode.v30.annotation.gtf  gencode.v30.GRCh38.genes.gtf

snakemake -j 999 --configfile config.yaml --use-conda --nolock --cluster-config cluster.json --cluster "sbatch -A {cluster.account} -p {cluster.partition}  -N {cluster.N} -n {cluster.n}  -t {cluster.time} --mem {cluster.mem}"