Do illumina instruments generate four fastq files per sample?
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2.6 years ago
ATCG ▴ 370

I have data for several treatments and replicates, I'm trying to understand the sample sheet and have once question: I have fastq files L001 L002 L003 L004 But it seems that all belong to the same sample. Do illumina instruments run each sample in 4 lanes and generate 4 fastq files for the same sample? Thanks for your time!

Fasq Illumina lane • 2.6k views
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Each file corresponds to data coming from a different lane. For example, the NextSeq has this pattern of L001 - L004.

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2.6 years ago
GenoMax 123k

Do illumina instruments run each sample in 4 lanes and generate 4 fastq files for the same sample?

Answer is it depends, on how the data was processed.

If the same sample (or pool) was run on more than one lane then the demultiplexing program (bcl2fastq) will generate files for each sample(s) that are lane specific (e.g. L001 = Lane1 etc). This behavior can be suppressed by using --no-lane-splitting option for bcl2fastq. This will generate a single file per sample containing all data from as many lanes the sample ran on.

Note 1: HiSeq flowcells can have 8 lanes and will produce as many files if same sample/pool ran on all lanes.
Note 2: Some Illumina sequencers have optically separate lanes but run the same pool across entire flowcell (NextSeq, NovaSeq without XP kit).