[vg giraffe] Unable to add read group to BAM file

Question

[saruman]

Hi everyone,

I am aligning short read sequences (HG002) against the Human pangenome reference graph using VG Giraffe. Specifically, I am using the .gbz and .hapl index files to generate the BAM file. The alignment works without issues; however, I am unable to add the read group (I can do it using Picard, though I would like to spare this additional step).

singularity exec \
    -B /fs/scratch/PAS2525/trimmed:/input \
    -B /fs/scratch/PAS2525/output:/output \
    -B /fs/scratch/PAS2525/References/hprc:/index \
    -B /fs/scratch/PAS2525/tmp:/tmp \
    /fs/scratch/PAS2525/Singularity/vg_v1.68.0.sif \
    vg giraffe \
    --progress --rescue-algorithm none --output-format bam --threads 44 \
    --read-group 'ID:1\tSM:HG002\tLB:lib1\tPL:illumina\tPU:unit1' \
    --gbz-name /index/hprc.gbz \
    --haplotype-name /index/hprc.hapl \
    --kff-name /output/HG002.kff \
    --fastq-in /input/HG002.NovaSeq.pcr-free.35x.trimmed_R1.fastq.gz \
    --fastq-in /input/HG002.NovaSeq.pcr-free.35x.trimmed_R2.fastq.gz \
    > /fs/scratch/PAS2525/HG002.bam

Does anyone know why I am unable to add the read group? I have also tried the following string without success:

--read-group '@RG\tID:1\tSM:HG002\tLB:lib1\tPL:illumina\tPU:unit1'

Thank you in advance for any help.

[Arup Ghosh]

Share the error message.

[saruman]

The read alignment does not produce any error. The BAM file does not include the read group. I need to run picard AddOrReplaceReadGroups to add the read group.

[saruman]

It seems that using the following format fixes the problem:

--read-group "1 SM:HG002 LB:lib1 PL:illumina PU:unit1"

As shown below:

@HD VN:1.5  SO:coordinate
@SQ SN:chr10    LN:134758134    M5:fbf3087282695f275d460157fc7864a5
@SQ SN:chr11    LN:135127769    M5:7f144a0b4800c341a4b4f1518838cc59
@SQ SN:chr12    LN:133324548    M5:5086cf368e83b9cefd7bb19ef0f40d3c
@SQ SN:chr13    LN:113566686    M5:6f9949b26d5ece38c6cdefd48a516907
@SQ SN:chr14    LN:101161492    M5:7ad36d43a1ab059e190ab918f27cb5bf
@SQ SN:chr15    LN:99753195 M5:914e0733351aaa9a4354ca8efb527401
@SQ SN:chr16    LN:96330374 M5:fcc04e14d89b12c757d56c177261e248
@SQ SN:chr17    LN:84276897 M5:8627043bd24255e9f356e0c99ece251f
@SQ SN:chr18    LN:80542538 M5:b6a85aabc31b2d57f707aa679a8612c5
@SQ SN:chr19    LN:61707364 M5:c2c74cb0832b34b9e8eebf4b744a88c1
@SQ SN:chr1 LN:248387328    M5:e469247288ceb332aee524caec92bb22
@SQ SN:chr20    LN:66210255 M5:f1a31f30d3da50893470ff47bf0b5a38
@SQ SN:chr21    LN:45090682 M5:760794bbbe965d4d1e3884bf09d1f306
@SQ SN:chr22    LN:51324926 M5:0068189b6b7c95e860cce0815662293f
@SQ SN:chr2 LN:242696752    M5:e5ff97665b6025191d4d7dfa2cec4cf0
@SQ SN:chr3 LN:201105948    M5:0012f8a55c07f0b61e199736105e7257
@SQ SN:chr4 LN:193574945    M5:b7e17a4f043aab5bc45cdfd5cccf9deb
@SQ SN:chr5 LN:182045439    M5:e069fd69dce480b348767fa9827671b4
@SQ SN:chr6 LN:172126628    M5:ae714983adbd673c7c84b0b1daa91f7e
@SQ SN:chr7 LN:160567428    M5:3ff4435e65cf8cdd69a3150d09dd649f
@SQ SN:chr8 LN:146259331    M5:29aeec58cb54d9132e2283839f9fcc9c
@SQ SN:chr9 LN:150617247    M5:419dad1fb58ae389ad8828536b5d78cd
@SQ SN:chrM LN:16569    M5:c68f52674c9fb33aef52dcf399755519
@SQ SN:chrX LN:154259566    M5:1f2616c4cfa30104517e44dd3c426453
@SQ SN:chrY LN:62460029 M5:dd7264df17e7e4a4dac5b0f1f19dcfe0
@RG ID:1 LB:lib1 SM:HG002 PL:illumina PU:unit1
@PG ID:0    PN:vg
@PG ID:samtools PN:samtools PP:0    VN:1.22.1   CL:samtools reheader --command sed -E "s/[A-Za-z0-9_]+#0+#//g" -