Finally to get the counts

Now with all the bam files, indexed bam files and the bed file made previously we can get read counts per transcript. The moment you are anticipating is not far off. Make sure the bam.bai files are all in the same working directory.

We use BEDtools v2.24.0, with the multiBamCov option.

#PBS -P project
#PBS -N Syzygium_counts
#PBS -l nodes=1:ppn=20
#PBS -l walltime=01:00:00
#PBS -l pmem=24gb
#PBS -e ./Syzygium_counts.txt
#PBS -M email@sydney.edu.au
#PBS -m abe

# Load modules
module load bedtools

# Working directory
cd /working directory location

# Run bedtools script
multiBamCov -q 30 -p -bams AP0_coordSorted.bam AP1_coordSorted.bam AP2_coordSorted.bam -bed Syzygium.BED > Syzygium_counts.txt

The output from this will be a large text file with the numbers of reads that aligned to each transcript.

TRINITY_DN4630_c0_g1_i1 0 2449 68 0 18 30
TRINITY_DN4630_c0_g2_i1 0 457 2 0 4 4 0
TRINITY_DN4679_c0_g1_i1 0 410 2 0 2 0 0
TRINITY_DN4674_c0_g1_i1 0 227 4 0 0 0 0
TRINITY_DN4609_c0_g1_i1 0 265 0 0 0 0 0
TRINITY_DN4651_c0_g1_i1 0 1030 8 0 4 4

which you can put into a spreadsheet and name the columns. And the columns are in the order that the bam files are placed in the script. Of course you would include all the bam files for all samples and all times in the one run using bedtools.

Gene ID start end AP0 AP1 AP2
TRINITY_DN4630_c0_g1_i1 0 2449 68 0 18
TRINITY_DN4630_c0_g2_i1 0 457 2 0 4
TRINITY_DN4679_c0_g1_i1 0 410 2 0 2
TRINITY_DN4674_c0_g1_i1 0 227 4 0 0
TRINITY_DN4609_c0_g1_i1 0 265 0 0 0
TRINITY_DN4651_c0_g1_i1 0 1030 8 0 4

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out /  Change )

Google photo

You are commenting using your Google account. Log Out /  Change )

Twitter picture

You are commenting using your Twitter account. Log Out /  Change )

Facebook photo

You are commenting using your Facebook account. Log Out /  Change )

Connecting to %s