Failed second index. Any chances to get some data?

[dep]

Dear colleagues!
We have conducted a HiSeq 2500 dual index paired end run using TruSeq SBS Kit v3 - HS (200-cycles) chemistry and TruSeq PE Cluster Kit v3 - cBot – HS cluster generation kit for libraries prepared with Nextera Rapid Capture Exome. We had 93-8-8-93 cycles, but it seems like we had not enough ICB for the first part of the run due to an error with ICB splitting. So, we have very degraded data for cycles 102-111, including cycles 102-109 for the Index 2 reads. The amount of Q30 nucleotides for Index 2 read is about 0,1%.

Is it worth trying to get any info for the index 2 reads out of such a run to demultiplex the samples?

I believe there are no chances, but I must use any possibility to minimize the losses.

[GenoMax]

Are you using bcl2fastq for demultiplexing? Have you looked at the "undetermined" pool files to see if the indexes in read 2 have recoverable data (you can overcome one N's in that read but not more).

[dep]

Thank you for the reply!

Yes, I use bcl2fastq and there are many more Ns in read 2. So, I guess its lost?

[GenoMax]

I was asking about the "tag 2" sequences. They should be present in undetermined file (but concatenated like tag1tag2, 1:N:0:CGACAACCCNNCCC) in the fastq read ID's. You will find the files in Undetermined_indices/Sample_lane* directories.

[dep]

Yes, I mean the same. Many Ns there.

[GenoMax]

It is likely that your samples share a common index 2 otherwise you may have been able to get some of them separated based on index 1. Best to write this run off.

[dep]

OK, thank you for assistance!