Can we concatenate two fastq files from same sample but different runs
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2.9 years ago
Payal ▴ 100

Hi,

I have two paired end file sets for a sample.

Forward: sample_R1_001.fastq.gz, sample_R1_002.fastq.gz

Reverse sample_R2_001.fastq.gz, sample_R2_002.fastq.gz

This is not a multilane case. They were two separate runs from the same sample aliquot!!

  1. Should I concat R1_001 and R1_002 fastq files ?
    1. Or should I run them as two separate pipelines?
    2. Or should I run them separately till alignment and then do BAM merge like for multilane samples?

Thanks, Payal

sequencing RNA-Seq • 2.7k views
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Whether a separate run or separate lane makes little difference, they are still technical replicates. They should not be merged.

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s/should not be/should be/

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But is this the case also for variant calling? I remember in GATK they take care for lane in their analysis (case of de-multiplexed).

However, if you are generating multiple libraries for each sample, and/or multiplexing samples within and/or across sequencing lanes, the data must be de-multiplexed before pre-processing, typically resulting in multiple sets of FASTQ files per sample all of which should have distinct read group IDs (RGID).

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There are no appreciable lane effects any more so they have nothing to adjust.

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Whether a separate run or separate lane makes little difference, they are still technical replicates. They should not be merged.

I have not seen any lane practical lane effects. Illumina offers a --no-lane-splitting option for their post processing software (bcl2fastq) which will make a single file per sample, if the same pool ran across multiple lanes of a flowcell.

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23 months ago
anamaria ▴ 30

If I am not wrong, --no-lane-splitting is only available for bcl2fastq2, not for the bcl2fastq.

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