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基于相对PCR扩增能力评估,开发从大麻树脂中提取DNA的快速简便方法。

Development of rapid and simple method for DNA extraction from cannabis resin based on the evaluation of relative PCR amplification ability.

作者信息

Yamamuro Tadashi, Iwata Yuko T, Segawa Hiroki, Kuwayama Kenji, Tsujikawa Kenji, Kanamori Tatsuyuki, Inoue Hiroyuki

机构信息

National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa, Chiba 277-0882, Japan.

National Research Institute of Police Science, 6-3-1 Kashiwanoha, Kashiwa, Chiba 277-0882, Japan.

出版信息

Forensic Sci Int. 2018 Jun;287:176-182. doi: 10.1016/j.forsciint.2018.03.044. Epub 2018 Apr 4.

Abstract

In recent years, analysis of cannabis DNA has been increasingly used in forensic drug tests. However, in the case of cannabis resin, a processed marijuana product, complicated procedures are required for the extraction of clean DNA, as the presence of various impurities inhibits PCR amplification. Therefore, in this study, we attempted to identify the factors that would allow quick and simple DNA extraction from cannabis resin with a commercially available kit. We also constructed a simple assay system for comparing relative amplification efficiencies by end-point PCR and used it to evaluate the purity of the obtained DNA solutions. For extraction with a kit that contains a silica column, reducing the starting amount of resin, using the residue remaining after methanol extraction, dilution of the final solution, extraction with an equal amount of powdered activated carbon or an excess amount of polyvinylpolypyrrolidone, and the addition of an appropriate amount of polyvinylpyrrolidone to the solution after extraction were effective measures that improved amplification efficiency. Furthermore, the use of the most rapid alkaline extraction kit combined with the addition of powdered activated carbon allowed obtaining DNA solutions with sufficient amplification efficiency in about 10min. These findings should be useful for routine DNA analysis of cannabis resin during forensic examination.

摘要

近年来,大麻DNA分析在法医毒品检测中越来越多地被使用。然而,对于大麻树脂这种经过加工的大麻制品,由于各种杂质的存在会抑制PCR扩增,因此提取纯净DNA需要复杂的程序。所以,在本研究中,我们试图确定能够使用市售试剂盒从大麻树脂中快速简单提取DNA的因素。我们还构建了一个简单的检测系统,通过终点PCR比较相对扩增效率,并用于评估所获得DNA溶液的纯度。对于使用含有硅胶柱的试剂盒进行提取,减少树脂的起始量、使用甲醇提取后剩余的残渣、稀释最终溶液、用等量的粉末状活性炭或过量的聚乙烯聚吡咯烷酮进行提取,以及提取后向溶液中加入适量的聚乙烯吡咯烷酮,都是提高扩增效率的有效措施。此外,使用最快的碱性提取试剂盒并加入粉末状活性炭,能够在约10分钟内获得具有足够扩增效率的DNA溶液。这些发现对于法医检查期间大麻树脂的常规DNA分析应该是有用的。

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