Quantitative intra-short interspersed element PCR for species-specific DNA identification
Quantitative intra-short interspersed element PCR for species-specific DNA identification
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DOI：
10.1016/s0003-2697(03)00095-2
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发表时间：
2003-05-15
影响因子：
2.9
通讯作者：
Batzer, MA
中科院分区：
文献类型：
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作者：
Walker, JA;Hughes, DA;Batzer, MA
我们设计并评估了四种基于短散布元素(SINES)的PCR扩增方法，用于牛、猪、鸡和反刍动物DNA的物种特异性检测和定量。为了应对牛海绵状脑病的流行，对这些类型的方法的需求急剧增加。在基于SYBR Green的检测中，最低有效定量水平分别为0.1、0.01、5和1pg的起始DNA模板，使用我们的牛、猪、鸡和反刍动物物种特异性的基于Sine的PCR分析。背景与来自其他14个物种的DNA模板的交叉扩增可以忽略不计。通过从混合(复杂)来源中准确检测痕量物种特异性DNA的能力，进一步证明了PCR扩增片段的物种特异性。在牛、猪和鸡的10 ng DNA模板混合物中，牛的DNA检测到0.005%(0.5pg)，猪的DNA检测到0.0005%(0.05pg)，鸡的DNA检测到0.05%(5pg)。我们还使用这些检测方法测试了六种商业购买的肉类产品。我们在这里报道的基于Sine的聚合酶链式反应方法是物种特异性的，具有高度的敏感性，将提高对来自这些物种的DNA序列的检测限。(C)2003年埃尔塞维尔科学公司(美国)。版权所有。
We have designed and evaluated four assays based upon PCR amplification of short interspersed elements (SINEs) for species-specific detection and quantitation of bovine, porcine, chicken, and ruminant DNA. The need for these types of approaches has increased drastically in response to the bovine spongiform encephalopathy epidemic. Using SYBR Green-based detection, the minimum effective quantitation levels were 0.1, 0.01, 5, and 1 pg of starting DNA template using our bovine, porcine, chicken, and ruminant species-specific SINE-based PCR assays, respectively. Background cross-amplification with DNA templates derived from 14 other species was negligible. Species specificity of the PCR amplicons was further demonstrated by the ability of the assays to accurately detect trace quantities of species-specific DNA from mixed (complex) sources. Bovine DNA was detected at 0.005% (0.5pg), porcine DNA was detected at 0.0005% (0.05pg), and chicken DNA was detected at 0.05% (5pg) in a 10-ng mixture of bovine, porcine, and chicken DNA templates. We also tested six commercially purchased meat products using these assays. The SINE-based PCR methods we report here are species-specific, are highly sensitive, and will improve the detection limits for DNA sequences derived from these species. (C) 2003 Elsevier Science (USA). All rights reserved.