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Old 07-27-2016, 03:48 AM   #1
Location: Spain

Join Date: Jun 2010
Posts: 56
Default Strange Illumina fastq header

Hi all,

I have received two paired end fastq files to analyze and when I look at the header I realized the following:

File 1
@HWI-ST1131:130302:D1RGUACXX:6:1101:1133:2071 1:N:0:

File 2
@HWI-ST1131:130302:D1RGUACXX:6:1101:1133:2071 3:N:0:
Illumina explains the header here:

and as I have always seen the third from last character tells you about the read number; 1 for single read or read 2 of paired-end. In this case I have a number 3 as you can see.

Has anyone seen this? Maybe it is a very common issue but I never saw it and I am suspicious about it.

Thanks in advance,

Last edited by chariko; 07-27-2016 at 03:56 AM.
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Old 07-27-2016, 03:54 AM   #2
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Location: East Coast USA

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Posts: 6,884

If the tag read is output in a separate file (which you can optionally do with bcl2fastq v.2.x) then you will get (for a 1D barcode run) following files:

R1 = Read 1
R2 = Tag/Barcode read
R3 = Read 2

Since some tools expect to see #2 in second file you may need to change that 3 to a 2 for all fastq headers.
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Old 07-27-2016, 03:59 AM   #3
Location: Spain

Join Date: Jun 2010
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Genomax Thank you very much for your response.
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fastq, header, illumina, read number

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