15重快速STR复合扩增体系的构建

发布时间：2018-07-04 15:08

本文选题：法医遗传学 + 短串联重复序列　；参考：《法医学杂志》2016年01期

【摘要】：目的建立一套15重快速STR复合扩增体系。方法选择14个常染色体基因座以及1个性别基因座,采用Fast Start Taq DNA聚合酶系统,以DNA标准品9947A为模板,通过筛选扩增条件、选择热启动酶用量、调整引物平衡、优化快速扩增程序、筛选反应缓冲液、选择反应体系以及筛选添加剂等一系列复合扩增实验,比较各条件下等位基因丢失和非特异性扩增情况。结果在以1 ng DNA为模板、0.4μL聚合酶及10×Fast Start高保真反应缓冲液构成10μL快速体系的条件下,32 min即可获得标准DNA全部15个STR基因座的完整分型,无等位基因丢失和非特异性扩增现象,等位基因均衡性良好。同时,5%甘油、0.01%明胶、0.05%明胶和5 mmol/L硫酸铵可作为PCR扩增过程中拟加入的反应添加剂。结论本研究建立的15重快速STR复合扩增体系可以明显缩短反应时间,提高样品检测效率。
[Abstract]:Objective to establish a 15-fold rapid STR multiplex amplification system. Methods 14 autosomal loci and 1 sex locus were selected. Fast start Taq DNA polymerase system was used, DNA standard strain 9947A was used as template, the amount of hot priming enzyme was selected and primer balance was adjusted by screening amplification conditions. A series of multiplex amplification experiments such as optimization of rapid amplification program, screening of reaction buffer, selection of reaction system and screening of additives were carried out to compare the allele loss and non-specific amplification under different conditions. Results the complete typing of all 15 STR loci of standard DNA could be obtained by using 1 ng DNA as template, 0.4 渭 L polymerase and 10 脳 Fast start high fidelity reaction buffer as 10 渭 L rapid reaction system. There was no allelic loss or nonspecific amplification. Allele equilibrium was good. At the same time, 5% glycerol, 0. 01% gelatin, 0. 05% gelatin and 5 mmol / L ammonium sulfate can be used as additives in PCR amplification. Conclusion the 15-step rapid STR multiplex amplification system can significantly shorten the reaction time and improve the detection efficiency.
【作者单位】：中国人民公安大学刑事科学技术学院;公安部物证鉴定中心北京市现场物证检验工程技术研究中心;公安部物证鉴定中心法医遗传学公安部重点实验室;清华大学医学系统生物学研究中心;
【基金】：中央级公益性科研院所基本科研业务费专项资金计划(2012JB012) 公安部技术研究计划(2014JSYJA011)
【分类号】：D919

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