Exome Seq Alignment - What to do with unpaired reads post trimming

Question

[andrew.j.skelton73]

Hi,

I have some paired end exome sequencing, and after checking out the samples in fastqc, I found there was an enrichment for Illumina adapters. I used Trimmomatic to remove the adapters (using Illumina Clip), which gives me paired reads, and unpaired reads for forward and reverse respectively post trimming.

I'm following the GATK best practises for variant calling, which recommends BWA MEM for alignment. I can use the paired, trimmed reads for to align, but there's quite a large proportion of reads which are trimmed and unpaired. The bedrock of my question is how can I use these effectively?

Would the best way be to do an alignment with the pairs, then unpaired, and combine the SAM files in some way? Or even merge the BAMs?

Anyone had a similar experience? Advice is welcome!

Thanks

Answer 1

[Brice Sarver]

I do this regularly with exome datasets.

Reads where one mate fails but are otherwise fine are included in a single-end file. It's still good data! Map them the same way you do the PE data, and subsequently combine the BAMs using Picard's MergeSamFiles or another approach.

Some cleaning approaches make this easy. I use expHTS to manage my cleaning now - does a fantastic job and is blazingly fast with stream-based processing. Documentation is forthcoming, but the developers are happy to help.