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Selective Lysis Enables Effective Whole Genome Bisulphite Sequencing of Buccal Epithelial Samples
EP23797
Poster Title: Selective Lysis Enables Effective Whole Genome Bisulphite Sequencing of Buccal Epithelial Samples
Submitted on 01 Feb 2016
Author(s): Andrew D. Johnston, Bassam El-Fahmawi, John M. Greally
Affiliations: Department of Genetics, Albert Einstein College of Medicine, MAWI DNA Technologies
This poster was presented at EPIGENOMICS 2016
Poster Views: 1,673
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Poster Information
Abstract: As population-based association studies increasethe number of individuals sampled to generate more statistical power, the selection of a cost-effective, easy-to-use, and time-sparing methodology for the collection and processing of samples becomes essential. With declining costs of sequencing, whole genome approaches are becoming more financially feasible. However, oral samples such as the mixed cell types of saliva or the homogeneous buccal epithelial cells from exfoliative brushing have significant amounts of microbial contamination, which dilutes sequencing results of the target human sample. We therefore explored the utility of using Mawi DNA Technologies’ buccal DNA collection kit, iSwab-DNA, since the storage buffer in the kit involves selective lysis of human cells, thus reducing microbial contamination. We tested its use for whole genome bisulphite sequencing (WGBS) – an assay requiring high coverage that would be sensitive to bacterial contamination.

We compared iSwab to our previously used DNA collection strategy based on the alcohol-based fixative PreserveCyt, and found that iSwab collected ~2-5x more DNA in 1/12 of the time, even after remaining at room temperature for weeks. Most importantly, the microbial contamination rates were much lower in iSwab-collected samples, especially when the collection occurred after eating (8% vs. 31%), allowing minimally contaminated iSwab samples to achieve a total efficiency (# of trimmed-mapped-deduplicated reads / # of raw reads) of ~70% and indicating that only ~3 lanes of Illumina HiSeq2500 would be necessary to reach 30x coverage. Furthermore, we examined the coverage profiles of three iSwab-sampled individuals and found uniform read coverage after accounting for GC-bias and mappability. Altogether, we find iSwab to be an ideal buccal epithelium collection method for studies wishing to apply downstream sequencing-based approaches.
Summary: As population-based association studies increase the number of individuals sampled to generate more statistical power, the selection of a cost-effective, easy-to-use, and time-sparing methodology for the collection and processing of samples becomes essential. Oral samples like the mixed cell types of saliva or the homogeneous buccal epithelial cells have significant amounts of microbial contamination, diluting sequencing results of the target human sample, which is unlike iSWAB-DNA samples. References: Berko, E. R., Suzuki, M., Beren, F., Lemetre, C., Alaimo, C. M., Calder, R. B., et al. (2014). Mosaic epigenetic dysregulation of ectodermal cells in autism
spectrum disorder. PLoSGenetics, 10(5), e1004402. http://doi.org/10.1371/journal.pgen.1004402
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