Zhang Lin, Xin Junping, Chen Guodi, Liao Miao, Li Ronghua
West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu, Sichuan, 610041 P. R. China.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2002 Feb;19(1):17-21.
To resolve the problem of the accuracy and standardization of short tandem repeat-polymerase chain reaction (STR-PCR) typing in forensic practice, the authors have designed a new method of producing standard D12S391 allelic ladder.
Nine different PCR amplified D12S391 allelic fragments were isolated from the gel, eluted into the distilled water and re-amplified by PCR. The purified allelic fragments were then blunt-end subcloned individually into the pUC plasmid vectors and transfected into competent E.coli DH5 alpha(TM) cells. The sequencing results confirmed that the size and the structure of the inserts were correct. The recombinant plasmids DNA with 9 inserts were then used as templates for PCR re-amplification to generate D12S391 standard ladder.
With the ladder, the authors studied the genetic polymorphisms of D12S391 locus in six populations (German, Japanese and Chinese south-western Han, northern Han, Weiwu'er and Hui populations), and the respective primary data in the six populations were obtained. D12S391 locus showed high polymorphism in all six populations, and its exclusion power and discrimination power are 0.609-0.786 and 0.940-0.952 respectively.
The results demonstrate that the standard ladder generated via this method is excellent, and D12S391 locus is robust for genetic research and forensic application.
为解决法医实践中短串联重复序列-聚合酶链反应（STR-PCR）分型的准确性和标准化问题，作者设计了一种制备标准D12S391等位基因阶梯的新方法。
从凝胶中分离出9个不同的PCR扩增D12S391等位基因片段，洗脱到蒸馏水中，再通过PCR重新扩增。然后将纯化的等位基因片段分别平端亚克隆到pUC质粒载体中，并转染到感受态大肠杆菌DH5α（TM）细胞中。测序结果证实插入片段的大小和结构正确。然后将带有9个插入片段的重组质粒DNA用作PCR重新扩增的模板，以生成D12S391标准阶梯。
利用该阶梯，作者研究了6个群体（德国、日本以及中国西南汉族、北方汉族、维吾尔族和回族群体）中D12S391基因座的遗传多态性，并获得了这6个群体各自的原始数据。D12S391基因座在所有6个群体中均表现出高度多态性，其排除能力和鉴别能力分别为0.609-0.786和0.940-0.952。
结果表明，通过该方法产生的标准阶梯效果良好，D12S391基因座在遗传研究和法医应用方面具有可靠性。
