Stangegaard Michael, Frøslev Tobias G, Frank-Hansen Rune, Hansen Anders J, Morling Niels
Section of Forensic Genetics, Department of Forensic Medicine, Faculty of Health Sciences, University of Copenhagen, Copenhagen, Denmark.
J Lab Autom. 2011 Apr;16(2):134-40. doi: 10.1016/j.jala.2010.11.003.
We have implemented and validated automated protocols for DNA extraction and PCR setup using a Tecan Freedom EVO liquid handler mounted with the Te-MagS magnetic separation device (Tecan, Männedorf, Switzerland). The protocols were validated for accredited forensic genetic work according to ISO 17025 using the Qiagen MagAttract DNA Mini M48 kit (Qiagen GmbH, Hilden, Germany) from fresh whole blood and blood from deceased individuals. The workflow was simplified by returning the DNA extracts to the original tubes minimizing the risk of misplacing samples. The tubes that originally contained the samples were washed with MilliQ water before the return of the DNA extracts. The PCR was setup in 96-well microtiter plates. The methods were validated for the kits: AmpFℓSTR Identifiler, SGM Plus and Yfiler (Applied Biosystems, Foster City, CA), GenePrint FFFL and PowerPlex Y (Promega, Madison, WI). The automated protocols allowed for extraction and addition of PCR master mix of 96 samples within 3.5h. In conclusion, we demonstrated that (1) DNA extraction with magnetic beads and (2) PCR setup for accredited, forensic genetic short tandem repeat typing can be implemented on a simple automated liquid handler leading to the reduction of manual work, and increased quality and throughput.
我们已经使用配备了Te-MagS磁分离装置（瑞士曼纳多夫的帝肯公司）的帝肯Freedom EVO液体处理仪，实施并验证了用于DNA提取和PCR设置的自动化方案。根据ISO 17025标准，使用Qiagen MagAttract DNA Mini M48试剂盒（德国希尔德的Qiagen有限公司），从新鲜全血和死者血液中提取DNA，对这些方案进行了认证法医基因工作的验证。通过将DNA提取物放回原始管中简化了工作流程，将样品放错位置的风险降至最低。在放回DNA提取物之前，用超纯水冲洗最初装有样品的管子。PCR在96孔微量滴定板中设置。对以下试剂盒的方法进行了验证：AmpFℓSTR Identifiler、SGM Plus和Yfiler（美国加利福尼亚州福斯特城的应用生物系统公司），GenePrint FFFL和PowerPlex Y（美国威斯康星州麦迪逊的Promega公司）。自动化方案允许在3.5小时内提取并添加96个样品的PCR预混液。总之，我们证明了（1）使用磁珠提取DNA和（2）为认证的法医基因短串联重复序列分型设置PCR，可以在简单的自动化液体处理仪上实现，从而减少人工操作，提高质量和通量。
