Hi,
I've got a problem with specific bam files, I've downloaded only a slice of them (one chromosome), this results in some MATE_NOT_FOUND error when checking my bams with:
gatk ValidateSamFile -I input.bam -M SUMMARY
Giving the error:
Error Type Count
ERROR:MATE_NOT_FOUND 1408
ERROR:MISMATCH_FLAG_MATE_NEG_STRAND 18551
ERROR:MISMATCH_FLAG_MATE_UNMAPPED 733
I have tried two solutions, both decreases the number of MATE_NOT_FOUND but there are still a few remaining, I don't understand how it is possible.
1. I've tried with samtools:
samtools view input.bam -f 0x1 -f 0x2 -b -o input.fixed.bam
It should only keep "read paired (0x1)" and "read mapped in proper pair (0x2)".
Now I have this output:
Error Type Count
ERROR:MATE_NOT_FOUND 81
ERROR:MISMATCH_FLAG_MATE_UNMAPPED 524
2. I've tried with PrintReads (GATK):
gatk PrintReads -I input.bam -o input.fixed.bam --read-filter PairedReadFilter
It should only keep reads that are properly paired:
Now I have this output:
Error Type Count
ERROR:MATE_NOT_FOUND 81
ERROR:MISMATCH_FLAG_MATE_UNMAPPED 524
Why they are not all removed?
I've tried the solution proposed by GenoMax but it didn't work for me.
After many attempts with the bams I finally solved the problem with a more radical solution by reverting the bams to fastq
samtools sort -n -O BAM -o sorted.bam input.bam
samtools fastq -1 output_1.fastq.gz -2 output_2.fastq.gz -0 /dev/null -s /dev/null -n sorted.bam
Then realign with bwa mem.
I think it is not the optimal solution as data loss could occur (BQSR data?) but at least it works.
I came across this same problem with a BAM file mapped to some unknown reference version and did something similar to @lacb. I followed this blog post from Heng Li using samtools 1.18 and bwa 0.7.17-r1188. This was also referenced in Realigning BAM files to new reference:
samtools collate -Oun128 in.bam | samtools fastq -OT RG,BC - \
| bwa mem -pt8 -CH <(samtools view -H in.bam|grep ^@RG) ref.fa - \
| samtools sort -@4 -m4g -o out.bam -
Assuming you have right tags you could try: How to remove un-paired reads from BAM after filtering? or use the solution provided in the same thread as an answer.
Log in to answer this question.
