Indexing

Image Reference:

STAR software is used to index the chosen reference genome assembly, Lee Assembly v1. This will be used as the reference genome to be aligned to the Raw Data for Glycine Max.

STAR Index Code

#!/bin/tcsh

#BSUB -J starindices_Portfolio #job name

#BSUB -n 10 #number of nodes

#BSUB -W 2:0 #time for job to complete

#BSUB -o starindices.out.%J #output file

#BSUB -e starindices.err.%J #error file

# For running star to generate genome index

# Run in working directory /share/bitcpt/S23/UnityID/Soy

# Must run this in working directory with subdirectory named starindices/

set STAR=/usr/local/usrapps/bitcpt/star/bin/STAR

set IN=/share/bitcpt/S23/referenceGenomes/Portfolios/Glycine_max_Lee_v1/

${STAR} --runThreadN 10 --runMode genomeGenerate --genomeSAindexNbases 12 --genomeDir

starindices/ --genomeFastaFiles ${IN}/glyma.Lee.gnm1.BXNC.genome_main.fna --sjdbGTFfile ${IN}/glyma.Lee.gnm1.ann1.6NZV.gene_models_main.AGAT.gtf --sjdbOverhang 100

Work Flow and Description of Code

Work Flow

The job script should be placed in the working directory following the path:

/share/bitcpt/S23/UnityID/Portfolio

To the run the job:

bsub <"Job name"

To check the job is running:

bjobs

The output files of the code will look like:

Description of Code

This command defines the variable STAR to be the path to the STAR software:

set STAR=/usr/local/usrapps/bitcpt/star/bin/STAR

This command defines the variable IN to be the path to the reference genome chosen:

set IN=/share/bitcpt/S23/referenceGenomes/Portfolios/Glycine_max_Lee_v1/

This sets the number of threads:

--runThreadN 10

This is telling STAR to index genome:

--runMode genomeGenerate

Integer length (bases) of the SA pre-indexing string:

--genomeSAindexNbases 12

(–genomeSAindexNbases must be scaled down to min(14, log2(GenomeLength)/2 - 1).)

This tells STAR where to put the output:

--genomeDir starindices/

This tells STAR where the genome assembly file is:

--genomeFastaFiles ${IN}/glyma.Lee.gnm1.BXNC.genome_main.fna

This tells STAR where the annotation file is:

--sjdbGTFfile ${IN}/glyma.Lee.gnm1.ann1.6NZV.gene_models_main.AGAT.gtf

Length of RNA-seq reads - 1. Although, in most cases default value of 100 works as well as the ideal value

--sjdbOverhang 100

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