Low % Phred Q30 for one of the lanes in HiSeq 4000 SE50 run

Question

[kspata]

Hi All,

I observed a discrepancy for Illumina HiSeq 4000 SE 50 run. The Lane 7 had very low Phred Quality % bases compared to other two lanes belonging to same sample set. The %Q30 was in 50's while for others its >90%. The libraries are CHIPseq libraries.

The lanes 5 and 6 which have samples from the same project show good quality phred scores.

I delmultiplexed the run again using --barcode-mismatches 0 but found no change in Phred Quality. When i tried using --barcode-mismacthes 2, it gave an error Barcode collision for barcodes: GGCTAC, AGTCAA

Is this a de-multiplexing error or a run error?

Thanks!!

Answer 1

[GenoMax]

This is likely an instrument hardware related error. Focus/flow control could be anything. You should contact Illumina tech support and have them do a remote check on the run/instrument. If you are not part of the sequencing core then you should ask them to do it.

it gave an error Barcode collision for barcodes: GGCTAC, AGTCAA

This bcl2fastq error just means that the index sequences you have on that lane do not allow two errors when demultiplexing since indexes will overlap between two samples.