Anthropology, University of Florida
Saturday Afternoon, Ballroom B
Our ongoing project in the Democratic Republic of Congo is to investigate support for an epigenetic mechanism that mediates the effects of maternal prenatal stress on newborn birth outcome. We are interested in determining the methylation status of CpG sites throughout the genome, at coding and non-coding regions. In order to determine the most cost-effective method, we conducted a pilot study to create indexed reduced representation bisulfite sequencing (RRBS) libraries and sequence them for methylation quantification. In an effort to systematically evaluate the yield and reproducibility of RRBS libraries, we produced and sequenced four technical replicates from a single human placental sample. Libraries were tagged and sequenced on a single lane of an Illumina GAIIx yielding over 40 million reads. These reads yielded coverage for ~1.5 million CpGs per library. However, after estimating a minimum necessary coverage of 20x for accurate epi-genotyping and identifying library overlaps, the number of informative sites was reduced to ~0.2 million per library. Scaling up to include the total 75 samples in our study would further reduce the number of informative CpG sites per library, or sample. Thus, we opted to switch to Illumina’s HumanMethylation450 Bead Chip. Preliminary analysis indicates that ~485,000 CpG sites were assayed for 72 samples, far exceeding what could be acquired from RRBS at a similar price. Further analyses are ongoing.
Funding was provided by NSF grant BCS 1231264, University of Florida (UF) Clinical and Translational Science Institute and UF Research Opportunity Seed Fund.