McMichael Gai L, Gibson Catherine S, O'Callaghan Michael E, Goldwater Paul N, Dekker Gustaaf A, Haan Eric A, MacLennan Alastair H
The University of Adelaide, Disciplines of Obstetrics and Gynaecology, Women's and Children's Hospital, Adelaide, Australia.
J Biomol Tech. 2009 Dec;20(5):232-5.
We sought a convenient and reliable method for collection of genetic material that is inexpensive and noninvasive and suitable for self-collection and mailing and a compatible, commercial DNA extraction protocol to meet quantitative and qualitative requirements for high-throughput single nucleotide polymorphism (SNP) multiplex analysis on an automated platform. Buccal swabs were collected from 34 individuals as part of a pilot study to test commercially available buccal swabs and DNA extraction kits. DNA was quantified on a spectrofluorometer with Picogreen dsDNA prior to testing the DNA integrity with predesigned SNP multiplex assays. Based on the pilot study results, the Catch-All swabs and Isohelix buccal DNA isolation kit were selected for our high-throughput application and extended to a further 1140 samples as part of a large cohort study. The average DNA yield in the pilot study (n=34) was 1.94 microg +/- 0.54 with a 94% genotyping pass rate. For the high-throughput application (n=1140), the average DNA yield was 2.44 microg +/- 1.74 with a >or=93% genotyping pass rate. The Catch-All buccal swabs are a convenient and cost-effective alternative to blood sampling. Combined with the Isohelix buccal DNA isolation kit, they provided DNA of sufficient quantity and quality for high-throughput SNP multiplex analysis.
我们寻求一种便捷可靠的遗传物质采集方法，该方法价格低廉、非侵入性，适用于自我采集和邮寄，同时还需要一种与之兼容的商业DNA提取方案，以满足在自动化平台上进行高通量单核苷酸多态性（SNP）多重分析的定量和定性要求。作为一项试点研究的一部分，我们从34名个体中采集了口腔拭子，以测试市售的口腔拭子和DNA提取试剂盒。在用预先设计的SNP多重分析检测DNA完整性之前，先用Picogreen双链DNA在荧光分光光度计上对DNA进行定量。根据试点研究结果，我们选择了全捕获拭子和Isohelix口腔DNA分离试剂盒用于高通量应用，并将其扩展到另外1140个样本，作为一项大型队列研究的一部分。试点研究（n = 34）中的平均DNA产量为1.94微克±0.54，基因分型通过率为94%。对于高通量应用（n = 1140），平均DNA产量为2.44微克±1.74，基因分型通过率≥93%。全捕获口腔拭子是一种方便且经济高效的采血替代方法。与Isohelix口腔DNA分离试剂盒相结合，它们提供了足够数量和质量的DNA用于高通量SNP多重分析。
