Build RG information for BWA

Question

[hpapoli]

Hello,

There are many posts about building the read group information but I haven't found a complete one yet. For a sample called, Sample_A, with a fastq header from sample_A.R1.fastq.gz, as follows:

@A00181:639:HNTFMDSX5:2:1101:1018:1000 1:N:0:ACACTAAG+TTATGGAT

Is this ReadGroup correct?

@RG\\tID:HNTFMDSX5.2\\tPL:ILLUMINA\\tLB:Sample_A\\tSM:Sample_A\\tPU:HNTFMDSX5.2.SampleA

HNTFMDSX5: Flowcell ID

2: Flowcell lane

I am mostly uncertain about the value for LB and the value for the third field of PU (PU= {FLOWCELL_BARCODE}.{LANE}.{SAMPLE_BARCODE})

Thanks so much for your help!

Answer 1

[Pierre Lindenbaum]

PL:Illumina must be uppercase to be compatible with gatk tools.

https://samtools.github.io/hts-specs/SAMv1.pdf

Platform/technology used to produce the reads. Valid values: CAPILLARY, DNBSEQ (MGI/BGI),
ELEMENT, HELICOS, ILLUMINA, IONTORRENT, LS454, ONT (Oxford Nanopore), PACBIO (Pacific Bio-
sciences), SINGULAR, SOLID, and ULTIMA. This field should be omitted when the technology is
not in this list (though the PM field may still be present in this case) or is unknown.

[hpapoli]

thanks very much! What about the ID and PU field? Is the PU field correct as it is written now? Thanks again!

[GenoMax]

https://gatk.broadinstitute.org/hc/en-us/articles/360035890671-Read-groups should explain how to use read groups. This is a GATK requirement. Probably not needed for most other software.