A bioinformatic filter for improved base-call accuracy and polymorphism detection using the Affymetrix GeneChip® whole-genome resequencing platform

Pandya, Gagan A.; Holmes, Michael H.; Sunkara, Sirisha; Sparks, Andrew; Bai, Yun; Verratti, Kathleen; Saeed, Kelly; Venepally, Pratap; Jarrahi, Behnam; Fleischmann, Robert D.; Peterson, Scott N.

The remaining SNP calls are next put through the footprint effect filter. The occurrence of a real SNP in a query sample results in a destabilizing effect on 25-mers in the immediate vicinity of the SNP. This artifact, called the 'footprint effect', is illustrated in Figure 4 . The locus on the resequencing chip at which the SNP occurs contains two probes that hybridize perfectly with the sample over the entire 25-base length of the forward-and reversecomplement probes (only the forward strand is shown in the figure). However, at adjacent loci on the chip, which represent base positions near the SNP base, there are no probes that hybridize perfectly with the sample DNA. This is because, in general, the chip design tiles probes based on a single reference sequence, which does not contain the SNP base. As a result, the probes on the chip that represent reference sequence positions within 12 bases of the SNP location will all contain at least a single-base mismatch with the sample DNA. This mismatch decreases the reference probe hybridization intensities and increases the likelihood that an alternate sequence from a second location in the sample DNA will hybridize more strongly to a non-reference probe pair. This results in a mixture of reference calls, SNP calls and no-calls at the loci within 12 base positions adjacent to a genuine SNP, with reference calls predominant. This effect is exacerbated when two genuine SNPs occur within the same 25-base window. The footprint effect, like the alternate homology effect, is expected to be more pronounced in the context of a whole-genome hybridization, because of the larger number of hybridization targets in the sample.