Question about Base calling for pacbio data

Question

[kamel]

I looked at some links of GATK on pacbio data but I can not understand that they are the parameters that must be used on -variant_index_parameter , -stand_emit_conf , -stand_call_conf during the base calling by haplotypecaller.

Also for VariantFiltration of SNP and Indel

-filterExpression 'QD || FS || MQ || HaplotypeScore || MappingQualityRankSum || ReadPosRankSum

Could you give me an example of the base call on pacbio data with the values of these parameters.

[William Rowell]

I'm not really sure exactly what you're asking, but I'm happy to try to help out here. Are you looking recommendations for GATK HaplotypeCaller and VariantFiltration parameters for PacBio data? What type of PacBio data are you using, and how was it generated?

[kamel]

I want to know what values should be used for these parameters on pacbio sequel data (these are whole genome data from a haploid fungus). For example on the illumina data I use the values below:

For Base calling

$   HaplotypeCaller -R Reference.fa -I sorted_marked.bam -ploidy 1 -ERC GVCF   --variant_index_type LINEAR --variant_index_parameter 128000 -stand_emit_conf 10 -stand_call_conf 30 -o raw_gVCF.vcf

For SNP filtration

$    VariantFiltration -R reference.fa -V raw-snps.vcf --filterExpression 'QD < 2.0 || FS > 60.0 || MQ < 40.0 || HaplotypeScore > 13.0 || MappingQualityRankSum < -12.5 || ReadPosRankSum < -8.0' --filterName “my_snp_filter” -o filtered_snps.vcf

For Indel Filtration

$   VariantFiltration -R reference.fa -V raw-indel.vcf --filterExpression 'QD < 2.0 || FS > 200.0 || ReadPosRankSum < -20.0' --filterName “my_indel_filter” -o filtered_indels.vcf

I have no idea about the values that should be used for these parameters on pacbio sequel data. Or do I use the same values?

[lakhujanivijay]

Hello kamel ,

Please use the formatting bar (especially the code option) to present your post better. I've done it for you this time.

Thank you!

[William Rowell]

Are these subreads, or CCS?

[kamel]

these are subreads generated by sequel

[William Rowell]

I have parameters for calling and filtering short variants with GATK HC on Q20+ CCS reads, but I haven't had much luck with raw subreads.