Command line explain
Entering edit mode
2.2 years ago
f1978hp ▴ 30

I see a command line used to variant calling but i don't understand a particular point. The command line is below.

samtools mpileup -C50 -d10000 -L1500 -uf REF_FASTA -I BEDFILE --output-tagsDP,DP4 -r {} Sample.BAM

and so,

bcftools call -mv > sample.{}.vcf

The part that i don't understand is the parameter -C50. The samtools documentation said:

 -C, --adjust-MQ INT     

Coefficient for downgrading mapping quality for reads containing excessive mismatches. Given a read with a phred-scaled probability q of being generated from the mapped position, the new mapping quality is about sqrt((INT-q)/INT)*INT. A zero value disables this functionality; if enabled, the recommended value for BWA is 50.

Why use this parameter to downgrading MQ if there isn't a filter step after it?

Thanks for clarification!

mpileup bcftools • 1.1k views
Entering edit mode

Hello f1978hp ,

as the documentation says, the mapping quality is downgraded for reads with excessive mismatches. Mapping quality is just one parameter for bcftools to decide if there is a variant or not. Decreasing the mapping the quality in this ways, leads to fewer false positive variant calls at all. In my experiences often there is no additional filter step needed.

fin swimmer

BTW: samtools mpileup is deprecated and should be replaced by bcftools mpileup.

Entering edit mode

Hello fin swimmer!

But, how bcftools leads to fewer false positive variant calls based on decreasing the mapping quality? For example, if i had a read with excessive mismatches, the parameter -C 50 will decrease your MQ so at what point (argument) in this case bcftools will use the new value of MQ of this read to decide if there is a variant or not?

Entering edit mode

During variant calling bcftools compares the average mapping quality of reads supporting a variant with those they don't. If there is a significant difference it will skip this variant. As we decrease the mapping quality before this difference will be much higher for reads with excessive mismatches.

BTW: There is no need to post your comment twice. I will delete one of them.

Entering edit mode

Thanks fin swimmer!

Do you know if there is a way to create a BAM file with these reads that skipped a variant because of a high difference in mapping quality? I would like to know better about this and appreciate a lot if you recommend a bibliography.

PS: Sorry about the duplication.


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