Quality assessment of buccal versus blood genomic DNA using the Affymetrix 500 K GeneChip.

Jessica G Woo,Daniel Woo,Joseph P Broderick,Ranjan Deka,Guangyun Sun,Lisa J Martin,Subbarao Indugula,Mary Haverbusch

doi:10.1186/1471-2156-8-79

Abstract

BackgroundWith the advent of genome-wide genotyping, the utility of stored buccal brushes for DNA extraction and genotyping has been questioned. We sought to describe the genomic DNA yield and concordance between stored buccal brushes and blood samples from the same individuals in the context of Affymetrix 500 K Human GeneChip genotyping.ResultsBuccal cytobrushes stored for ~7 years at -80°C prior to extraction yielded sufficient double stranded DNA (dsDNA) to be successfully genotyped on the Affymetrix ~262 K NspI chip, with yields between 536 and 1047 ng dsDNA. Using the BRLMM algorithm, genotyping call rates for blood samples averaged 98.4%, and for buccal samples averaged 97.8%. Matched blood samples exhibited 99.2% concordance, while matched blood and buccal samples exhibited 98.8% concordance.ConclusionBuccal cytobrushes stored long-term result in sufficient dsDNA concentrations to achieve high genotyping call rates and concordance with stored blood samples in the context of Affymetrix 500 K SNP genotyping. Thus, given high-quality collection and storage protocols, it is possible to use stored buccal cytobrush samples for genome-wide association studies.

Highlights

With the advent of genome-wide genotyping, the utility of stored buccal brushes for DNA extraction and genotyping has been questioned
Concerns regarding the use of buccal brushes have included the lower quantity of genomic DNA isolated [4], lower quality of DNA [4,5], and the fidelity of results from buccal brushes compared with blood samples [5,6,7]
The current study suggests several methodologic recommendations for using buccal cytobrushes for genomewide association studies or similar large-scale SNP genotyping

Summary

Introduction

With the advent of genome-wide genotyping, the utility of stored buccal brushes for DNA extraction and genotyping has been questioned. We sought to describe the genomic DNA yield and concordance between stored buccal brushes and blood samples from the same individuals in the context of Affymetrix 500 K Human GeneChip genotyping. Concerns regarding the use of buccal brushes have included the lower quantity of genomic DNA isolated [4], lower quality of DNA [4,5], and the fidelity of results from buccal brushes compared with blood samples [5,6,7]. The Affymetrix 500 K GeneChip requires only 250 ng of total genomic DNA per chip, 500 ng total, and this DNA quantity has not changed with the recent release of the Affymetrix 5.0 and 6.0 chips, which enable genotyping up to 1.8 million genetic markers [9,10,11]. As the Affymetrix system uses restriction enzymes to recognize a particular double-stranded DNA (dsDNA) sequence, we elected to examine the dsDNA yield from buccal cytobrushes, rather than total DNA including both single and double stranded DNA

Methods

Results

Discussion

Conclusion