Troubleshooting Tips – bcl2fastq creates duplicate reads

Hi,

I have seen a few times where bcl2fastq (v2.20) will produce duplicate FASTQ entries in sequencing read IDs, raw sequences, & quality scores. This causes issues with downstreams tools like Picard MarkDuplicates (e.g. Exception in thread "main" htsjdk.samtools.SAMException: Value was put into PairInfoMap more than once).

I’m hoping to get some community input for how to troubleshooot this. For me, I did not get this issue when I removed the --no-lane-splitting option from my command and I checked the FASTQs to verify that one of the two identical reads had been removed (made sure to check across all L00X reads).

So far, I’ve also heard verifying the latest bcl2fastq version and re-running bcl2fastq as options (seqanswers link) – does anyone have any more methods or want to share their experience? Thanks in advance