Tutorial - Start diploid mapping for Day 2

Prepare for Day 2

To prepare for Day 2, try to start either BWA or Bowtie2 mapping of these two fastq files:

The data

$BI/ngs_course/human_variation/allseqs_R1.fastq
$BI/ngs_course/human_variation/allseqs_R2.fastq

against this reference:

The reference

$BI/ngs_course/human_variation/ref/hs37d5.fa

followed by converting the output to bam format, sorting the bam file, and indexing the output.

Hint

For either BWA or Bowtie, you need an indexed reference generated by either bwa index or bowtie-build. Perhaps one already exists? You then call the alignment program(s) bwa aln/bwa sampe or bowtie. Conversion, sorting, and indexing of the output are all done via samtools.

Solution

Submit to the TACC queue

Move into your scratch directory and create a new directory:

cds
mkdir day2
cd day2

Then create a commands file with the commands below and use launcher_creator.py followed by qsub to run them.

Mapping

For BWA, the commands are:

ONE LINE command for ALL bwa operations

module load bwa; bwa aln $BI/ngs_course/human_variation/ref/hs37d5.fa $BI/ngs_course/human_variation/allseqs_R1.fastq > r1.sai && bwa aln $BI/ngs_course/human_variation/ref/hs37d5.fa $BI/ngs_course/human_variation/allseqs_R2.fastq > r2.sai && bwa sampe $BI/ngs_course/human_variation/ref/hs37d5.fa r1.sai r2.sai $BI/ngs_course/human_variation/allseqs_R1.fastq $BI/ngs_course/human_variation/allseqs_R1.fastq > test.sam

For bowtie2:

One-line command for bowtie

module load bowtie/2.0.0b6; bowtie2 -t -x $BI/ngs_course/human_variation/ref/hs37d5_bowtie2 -1 $BI/ngs_course/human_variation/allseqs_R1.fastq -2 $BI/ngs_course/human_variation/allseqs_R2.fastq -S test.sam

Convert, sort, and index output:

One-line command for samtools

samtools view -S -b test.sam > test.bam && samtools sort test.bam test.sorted && samtools index test.sorted.bam

For a bit of extra challenge, you could make two separate commands files, one for each mapper, qsub them. Watch out - you'll need to make sure they both write output to different files, or run in different directories!

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Prepare for Day 2