can I map multiple (>2) fastq reads against and assembly using bwa?

Question

[sa91]

Hi

I want to map the output of 4 separate illumina smrt cells against a pacbio reference genome using BWA. I've indexed the full genome.

I can try run the the equivalent of this line, but using additional reads:

bwa mem ref.fa read1.fq read2.fq > aln-pe.sam

However, if I try

bwa mem ref.fa read1.fq read2.fq read3.fq read4.fq > aln-pe.sam

include read3.fq or more nothing runs.. Instead I get the usage instructions for bwa.

Is there a way to map 4 illumina runs against the reference genome at once, or am I misunderstanding this process?

There are simply 4 runs to increase robustness.

[ATpoint]

It is unclear what you do. Do you have different paired-end samples, all with R1 and R2? Should all the four samples go to the same SAM file or to different ones?

[sa91]

These are 4 separate runs of illumina output that I am supposed to analyse and compare to my pacbio genome that I sequenced. I want all 4 in the same SAM file.

[GenoMax]

4 separate illumina smrt cells against a pacbio reference genome using BWA

You are mixing terminologies here. PacBio sequencer uses SMRTCells where as Illumina sequencers use flowcells. Do you have 4 separate samples/lanes/flowcells worth Illumina data? Depending on the answer there would be a different solution.

[sa91]

Thanks for correcting me. I was getting confused and using the wrong terminology. It is from flow cells. I've been asked to map illumina short reads against my pacbio genome assembly to examine the coverage.

Answer 1

[swbarnes2]

Your first command lined is for paired end sequencing. Is that what you have?

If you do not have paired end sequencing, just two fastqs of the same single-end library, then your first command line is wrong.

If you have 4 fastqs that are all the same single end library, you can cat them all together before giving them to bwa

cat read*.fq | bwa mem ref.fa - > out.sam

[sa91]

Thank you! I didn't realise that command was specifically for paired end sequencing. That was where i went wrong. i'll cat the other fastqs together

Answer 2

[gayachit]

Hi There are some posts similar to yours. Have a look.

bwa for multiple fastq files , BWA mem on multiple samples, perl script for BWA-mem on multiple different files