How To Call Snp And Indel Based On A Sanger Sequences Based Bac Contig Vs Reference?
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9.1 years ago
William ★ 5.1k

I have BAC contig, assembled from sanger sequences, that I want to use for quality control on my short read based SNP and indel call's.

Which tool can I use to align the BAC sequence against the reference, and which tool do I then use to call SNP and Indel?

Or is it smarter just to use the Sanger sequences directly? To align them with BWA-mem and have GATK (UG or HC) call SNP and Indel?

sanger snp indel qualitycontrol • 4.8k views
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9.1 years ago
Mchimich ▴ 300

Hi You can use DNAdiff of MuMmer tool. It will detect for you SNPs, Deletions, insertions and other type of structural variation as inversions, trans-locations...etc. DNAdiff compare tow DNA sequences (Your BAC sequence with a reference) but unfortunatly it can not consider the quality of your DNA sequence as other tools do for NGS data. Please have a look to this manual http://nebc.nerc.ac.uk/bioinformatics/documentation/mummer/dnadiff.README. You have to donlowd and install MuMmer and after run DNAdiff with your BAC sequences and the reference it works very well !. Hope that will help

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It doesn't seem to work for a large reference genome (as I have) :

MUMmer3.23/mummer: suffix tree construction failed: textlen=2909701677 larger than maximal textlen=536870908

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9.1 years ago

I guess you could use BWA MEM and samtools?

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9.1 years ago
William ★ 5.1k

I aligned the BAC contig with bwa mem and used GATK UG to call SNP and Indels . The mapped reads must have a mapping quality above some treshold, mapping quality of 6 was to low. After I increased the minimum seed lenght to 400 for bwa mem my mapping quality was at 60 and GATK UG would call SNPs. To have the GATK UG call Indels on just the aligned BAC contig you need to add some extra parameters:

glm BOTH -minIndelCnt 1 -indelGOP 80 -stand_call_conf 0 -stand_emit_conf 0


The last part of the answer I got from the GATK forum: http://gatkforums.broadinstitute.org/discussion/comment/7086#Comment_7086