Indexing

#1-iv. Indexing

###Create a bowtie2 index

Create a bowtie index for chr22 and the ERCC spike-in sequences and write it to a 'bwt' sub-directory:

cd $RNA_HOME/refs/hg19/
mkdir -p bwt/chr22_ERCC92
bowtie2-build fasta/chr22_ERCC92/chr22_ERCC92.fa bwt/chr22_ERCC92/chr22_ERCC92
cp $RNA_HOME/refs/hg19/fasta/chr22_ERCC92/*.fa $RNA_HOME/refs/hg19/bwt/chr22_ERCC92/
ls bwt/chr22_ERCC92/

To create an index for all chromosomes instead of just chr22 you would do the following:

#cd $RNA_HOME/refs/hg19/
#mkdir bwt
#bowtie2-build fasta/hg19.fa bwt/hg19
#mv $RNA_HOME/refs/hg19/fasta/hg19.fa $RNA_HOME/refs/hg19/bwt/

---

###OPTIONAL ALTERNATIVE - STAR indexes Create index files for use with STAR

cd $RNA_HOME/refs/hg19/
mkdir -p star/chr22_ERCC92
STAR --runMode genomeGenerate --genomeDir $RNA_HOME/refs/hg19/star/chr22_ERCC92 --genomeFastaFiles $RNA_HOME/refs/hg19/fasta/chr22_ERCC92/chr22_ERCC92.fa --runThreadN 8 --sjdbGTFfile $RNA_HOME/refs/hg19/genes/genes_chr22_ERCC92.gtf --sjdbOverhang 100

####END OF OPTIONAL ALTERNATIVE - STAR indexes

####OPTIONAL ALTERNATIVE - HISAT2 indexes Create reference and splice site index files for use with HISAT2

cd $RNA_HOME/refs/hg19/
mkdir -p hisat2/chr22_ERCC92
hisat2-build $RNA_HOME/refs/hg19/fasta/chr22_ERCC92/chr22_ERCC92.fa $RNA_HOME/refs/hg19/hisat2/chr22_ERCC92/chr22_ERCC92

extract_splice_sites.py $RNA_HOME/refs/hg19/genes/genes_chr22_ERCC92.gtf > $RNA_HOME/refs/hg19/hisat2/chr22_ERCC92/splicesites.txt

####END OF OPTIONAL ALTERNATIVE - HISAT2 indexes

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