No index in the undetermined fastq file
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Entering edit mode
3 months ago
1769mkc ★ 1.3k

The data was generated in MiSeq , demux was done in LRM but it didn't yeild any data assigned to the sample, but the other issue is in the undermined files we are not able to see in any index in the header of the undermined fastq files, which seems bit strange.

Any suggestion how to troubleshoot ,since SAV looks all fine the index read have good Q30 scores as adding the information index read info

zcat Undetermined_S0_L001_R1_001.fastq.gz | head @M06594:100:000000000-DN78N:1:1101:16087:1338 1:N:0:0 TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTNTNNNNNTTTTTTTTTNNNNNNNNTTTTTTTTNNTTTTTTTTTTTT + 1111>1>10000AA//AA/>///////<<<</<//-------:-9-9-9-999-9--9-9-9-9-------9-9-999-9-9--9------9-----99--9---#:#####99--9----########--999---##9----9----99 @M06594:100:000000000-DN78N:1:1101:15529:1349 1:N:0:0 TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTCTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTNNNNNNNNTTTTTTTTTTTTTTTTTTTTTT + 11111>1>>>>0///AA/////>////<<//////--<<-----9-9--9---9----99----9-999--999-------99//999:-9-99-------999----;---9--------########-----;---99-----9---9- @M06594:100:000000000-DN78N:1:1101:15215:1371 1:N:0:0

Any suggestion how to troubleshoot, what could be the issue?

bcl2fastq • 930 views
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Entering edit mode
3 months ago
GenoMax 153k

we are not able to see in any index in the header of the undermined fastq files, which seems bit strange.

If phiX was spiked in then that is not indexed. So the undetermined reads can at least partly be phiX. Rest could be "junk" data as they all appear to be poly-T in example you show.

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yes 15% phix was spiked in, now since that was in LRM i did try in bcl2fastq i can see the index in the header but no difference in the output . I did try swapping the index, then Reverse compliment the i5 index but no difference as such.

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Entering edit mode
3 months ago

Run bcl2fastq or bcl-convert yourself, and the indices might turn up.

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that what i did it was a samplesheet issue it was resolved it seems in the LRM the samplesheet which was used having some issue.

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