sam_hrecs_error Malformed key:value pair

Question

[Ric]

Hi, I am using the following bash script to map all FASTQ files in a folder with Bowtie2 and should convert it to bam files:

#!/bin/bash
#usage: sh bowtie2_pairend_pbs.sh /data ref.fasta bams

mkdir $3

for r1 in $(find $1 -name "*_R1*.fq");
do
  output=$(basename $(echo $r1 | sed 's/_R1//g'))
  r2=$(echo $r1 | sed 's/_R1/_R2/g')

  #cat <<EOF
  qsub <<EOF
#!/bin/bash -l

#PBS -N $output
#PBS -l walltime=24:00:00
#PBS -j oe
#PBS -l mem=30G
#PBS -l ncpus=8
##PBS -m bea

cd \$PBS_O_WORKDIR

conda activate bowtie2

# Alignment
bowtie2 -p 8 \
--rg-id ${output} --rg ${output} --rg ${output} --rg ${output} --rg ${output} \
-x ${2%.*} -1 $r1 -2 $r2 | samtools sort -@ 8 - -o ${3}/${output}-bowtie2.sorted.bam

samtools index ${3}/${output}-bowtie2.sorted.bam
samtools stats ${3}/${output}-bowtie2.sorted.bam > ${3}/${output}-bowtie2.sorted.bam.stats
conda deactivate

EOF

done

Unfortunately, I got the following error

[E::sam_hrecs_error] Malformed key:value pair at line 6494: "@RG        ID:faspt-rm_adapters_correction.232.fq  faspt-rm_adapters_correction.232.fq     faspt-rm_adapters_correction.232.fq     faspt-rm_adapters_correction.232.fq     faspt-rm_adapters_correction.232.fq"
[E::sam_hrecs_error] Malformed key:value pair at line 6494: "@RG        ID:faspt-rm_adapters_correction.232.fq  faspt-rm_adapters_correction.232.fq     faspt-rm_adapters_correction.232.fq     faspt-rm_adapters_correction.232.fq     faspt-rm_adapters_correction.232.fq"
samtools sort: failed to change sort order header to 'coordinate'

How is it possible to fix it?

Thank you in advance

Answer 1

[jilguero888]

I had this error with samtools sort because I set wrongly the @RG header line when calling bowtie2, in particular because I did not specify the SM tag when adding text to the @RG header line with --rg. For example

wrong: --rg-id idxx --rg text
right: --rg-id idxx --rg SM:text

It worked to me.