The 82nd Annual Meeting of the American Association of Physical Anthropologists (2013)


Y-chromosome library construction for next-generation sequencing

SHANNON M. MCNULTY1, AIDA T. MIRĂ“-HERRANS2,3 and CONNIE J. MULLIGAN2.

1College of Agricultural and Life Sciences, University of Florida, Gainesville, 2Department of Anthropology, University of Florida, Gainesville, 3Genetics and Genomics Graduate Program, University of Florida, Gainesville

Thursday All day, Park Concourse Add to calendar

The Y-chromosome is particularly useful for population history inferences through the investigation of haplogroup diversity among males. These haplogroups are defined by single nucleotide polymorphisms (SNPs). The cost of high-throughput sequencing of the entire Y-chromosome of multiple individuals for population analyses renders it prohibitive for SNP investigation. Selectively enriching for those areas of the Y-chromosome that provide the most useful information, via library construction, can make next-generation sequencing cost-effective. To this end, we are creating a protocol in which we enrich for Y-chromosome fragments containing lineage-defining SNPs through hybridization with PCR products of selected haplogroups. Two thousand base pair fragments were generated from these PCRs to serve as probes to capture complementary sequences on the Y-chromosome. The resulting PCR products were sheared to create a tiling effect that increased sequence diversity and coverage during hybridization. Individual samples were tagged with unique identifying sequences, allowing multiple samples to be pooled and sequenced on a single lane of the Illumina GAIIx after capture on custom probes. Sixty-three primer pairs were designed to produce probes containing over 150 Y-chromosome SNPs. SNPs were chosen that define major haplogroups, as well as more divergent subgroups of E and J haplogroups, which are of particular interest in our samples. Ninety-five samples were uniquely tagged and pooled for library construction. Approximately 1.824 billion bases mapped to the Y-chromosome, allowing for haplogroup determinations of all 95 samples. This method provides an unprecedented amount of Y-chromosome sequence data in order to address questions that have been intractable thus far.

Samples sequenced in the study were collected with support of NSF grant BCS-0518530

Tweet
comments powered by Disqus