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利用同步拖拽系数改变(SCODA)技术有效去除提取 DNA 样本中的共纯化抑制剂。

Effective removal of co-purified inhibitors from extracted DNA samples using synchronous coefficient of drag alteration (SCODA) technology.

机构信息

Institute of Applied Genetics, Department of Forensic and Investigative Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, USA.

出版信息

Int J Legal Med. 2013 Jul;127(4):749-55. doi: 10.1007/s00414-012-0810-7. Epub 2012 Dec 20.

Abstract

Various types of biological samples present challenges for extraction of DNA suitable for subsequent molecular analyses. Commonly used extraction methods, such as silica membrane columns and phenol-chloroform, while highly successful may still fail to provide a sufficiently pure DNA extract with some samples. Synchronous coefficient of drag alteration (SCODA), implemented in Boreal Genomics' Aurora Nucleic Acid Extraction System (Boreal Genomics, Vancouver, BC), is a new technology that offers the potential to remove inhibitors effectively while simultaneously concentrating DNA. In this initial study, SCODA was tested for its ability to remove various concentrations of forensically and medically relevant polymerase chain reaction (PCR) inhibitors naturally found in tissue, hair, blood, plant, and soil samples. SCODA was used to purify and concentrate DNA from intentionally contaminated DNA samples containing known concentrations of hematin, humic acid, melanin, and tannic acid. The internal positive control (IPC) provided in the Quantifiler™ Human DNA Quantification Kit (Life Technologies, Foster City, CA) and short tandem repeat (STR) profiling (AmpFℓSTR® Identifiler® Plus PCR Amplification Kit; Life Technologies, Foster City, CA) were used to measure inhibition effects and hence purification. SCODA methodology yielded overall higher efficiency of purification of highly contaminated samples compared with the QIAquick® PCR Purification Kit (Qiagen, Valencia, CA). SCODA-purified DNA yielded no cycle shift of the IPC for each sample and yielded greater allele percentage recovery and relative fluorescence unit values compared with the QIAquick® purification method. The Aurora provided an automated, minimal-step approach to successfully remove inhibitors and concentrate DNA from challenged samples.

摘要

各种类型的生物样本在提取适合后续分子分析的 DNA 时都存在挑战。常用的提取方法,如硅胶膜柱和酚氯仿,虽然非常成功,但对于某些样本,仍可能无法提供足够纯净的 DNA 提取物。同步拖曳系数改变(SCODA),应用于博奥基因组公司的极光核酸提取系统(Boreal Genomics,温哥华,BC),是一种新技术,它有可能有效地去除抑制剂,同时浓缩 DNA。在这项初步研究中,SCODA 被测试了其去除法医和医学相关聚合酶链反应(PCR)抑制剂的能力,这些抑制剂自然存在于组织、毛发、血液、植物和土壤样本中。SCODA 被用于从故意污染的 DNA 样本中纯化和浓缩 DNA,这些样本中含有已知浓度的血红素、腐殖酸、黑色素和单宁酸。Quantifiler™ Human DNA Quantification Kit(Life Technologies,福斯特城,CA)中的内部阳性对照(IPC)和短串联重复(STR)分析(AmpFℓSTR®Identifiler®Plus PCR 扩增试剂盒;Life Technologies,福斯特城,CA)用于测量抑制效果,从而评估纯化效果。与 QIAquick®PCR 纯化试剂盒(Qiagen,瓦伦西亚,CA)相比,SCODA 方法总体上提高了高度污染样本的纯化效率。与 QIAquick®纯化方法相比,SCODA 纯化的 DNA 对每个样本的 IPC 没有循环位移,并且具有更高的等位基因百分比恢复和相对荧光单位值。Aurora 提供了一种自动化的、最小步骤的方法,成功地从受挑战的样本中去除抑制剂并浓缩 DNA。

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