Kun:LabNotes/ExonomeSeq/2009-11-15
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Analysis of iPS exome sequencing data
- Data used:
- Libraries were constructed with the 1-adaptor protocol.
- DF6/Foreskin: HL022, two lanes per line, captured with probe set #1-7;
- CV-iPS/CV-fibroblast:
- HL020, two lanes per line, captured with probe set #8-9;
- HL022, three lanes for CV-iPS, captured with probe set #1-7;
- Read mapping and variant calling.
- Reads that contain potential H1/H2 capturing arms are identified and removed by mapping to a database that contains H1/H2 with 20bp flanking sequences.
- The remaining reads were mapped, down-sampled and fed to Samtools.
- This is the current mapping script: variantCallerBowtieSam.pl.
- Potential mutations were identified by comparing two pileup files ( pileup2variantsPair.pl);
- To be considered as a candidate mutation, I required that no single read contains the second allele in the reference sample.
- Results:
- 4 candidates of single nucleotide changes (SNCs) found in CV-iPS:CV-iPS_vs_fibro_set8-9_HL020;
- 5 candidates of SNCs found in DF6-9-9:DF6_Foreskin_HL022;
- Nine pairs of PCR primers were designed for these variants. However, ZNF708-K259R will be excluded in the first validation experiment because the flanking sequence has many highly similar homologs on the same chromosome.