混合DNA分型结果的解释及法医学应用

发布时间：2018-06-08 07:22

本文选题：混合DNA-STR分型 + 限制性组合分析法　；参考：《山西医科大学》2009年硕士论文

【摘要】： 目的:通过制备不同混合比例的模拟混合DNA,利用混合DNA分型中的峰高或峰面积信息,采用一定的参数标准对建立在峰高或峰面积基础上各基因座的表型组合进行分析,筛选出合理的表型组合,建立限制性组合分析法对混合DNA分型的结果进行解释,从而对男/女两人混合DNA分型的组分来源进行认定。并通过实际案例的验证,对建立的分析法在法医学应用中的适用范围进行探索,为解决法医学应用中男/女两人混合DNA分型结果的解释提供科学合理的途径。方法:选择AB-YfilerTM试剂盒中007阳性对照品作为已知男性DNA,选择AB-IdentifilerTM试剂盒中9947A阳性对照品作为已知女性DNA(两者浓度均为0.1ng/μl)。制备8个梯度比例的25例常量和不同混合比例不同混合DNA总模板量的19例低拷贝男/女两人模拟混合DNA(每个梯度比例同时做三次平行试验),应用杂合型均衡比和混合比两个参数,通过性别基因座估计男/女两人混合DNA中男性DNA所占的比例,对在峰高(峰面积)信息基础上各基因座的表型组合进行限制性的分析,筛选出合理的表型组合,建立限制性组合分析法。对男性组分混合比为95%、50%、5%的9个模拟混合样本的峰高和峰面积信息分别进行分析,比较其在限制性组合分析法中的应用有无差别。通过12例常量和5例低拷贝男/女两人混合DNA案件对建立的分析法进行验证,观察分析结果。对混合DNA分型中出现的一个等位基因缺失、两个等位基因缺失及低拷贝DNA分型中等位基因缺失三种类型,参考Peter Gill推荐的相应的量化公式[2]进行似然率(LR)计算。结果:25例模拟常量混合DNA分析的结果显示:3例混合样本检出的基因座数为15个(检出率100%),5例检出数为14个(检出率93%),4例检出数为13个(检出率87%), 6例检出数为12个(检出率80%),3例检出数为11个(检出率73%),最低检出数为8个(检出率53%)仅1例;在男性组分混合比为95%的三次重复混合样本中,分离得到的男性DNA分型依次出现2个、1个和4个等位基因缺失,其余混合样本均无缺失;6例混合样本出现等位基因的错判,其中5例样本错判数1个基因座、1例错判2个基因座,6例中的3例均为男性组分混合比为30%的样本;25例混合样本的似然率(LR)值均达到14个数量级以上。19例模拟低拷贝混合DNA分析的结果显示:1例低拷贝混合样本的检出基因座数为12个(检出率80%),1例检出数为11个(检出率73%),最低检出基因座数为3个(检出率20%)共4例;6例样本出现等位基因缺失,缺失集中在男性混合比低于20%或混合DNA模板量为100pg的样本中;18例样本出现等位基因错判;同一混合比的三次重复样本间LR值差别多数在6个数量级以内。选择峰高与峰面积信息进行混合DNA的分析在检出基因座数、等位基因缺失数和错判基因座数三方面无明显差别。 17例混合DNA案件验证的结果显示:12例常量混合DNA案件中,4例案件检出基因座数为15个,2例检出数为14个,1例检出数为13个,2例检出数为12个,2例检出数为11个,最低检出数为10个仅l例;5例案件出现等位基因缺失,其中4例均缺失1个基因座、1例缺失3个基因座;12例案件均无等位基因错判。5例低拷贝混合DNA案件中,最高检出基因座数为9个,最低检出数仅为3个;仅1例出现缺失现象;错判基因座数共10个。结论:结合44例模拟混合DNA和17例混合DNA案件的分析,建立的限制性组合分析法的适用范围是:混合DNA分型的15个基因座中最高等位基因条带的相对荧光强度应大于1000rfu,且混合比大于30%的男/女两人混合DNA。峰高或峰面积信息均可进行分析。在低拷贝混合DNA分析中,由于等位基因错判现象的发生无法进行量化计算,该分析法在此应用受到限制。
[Abstract]:Objective : To establish a reasonable phenotypic combination by using the peak height or peak area information in the mixed DNA typing by preparing the mixed DNA with different mixing ratios . The results of the typing of mixed DNA based on the peak height or peak area were analyzed by using a certain parameter standard . The application of the established analytical method in forensic medical application was explored through the verification of practical cases . The method provided scientific and reasonable way for solving the interpretation of the mixed DNA typing of male / female in forensic application .

Methods : In the AB - YfilerTM kit , 007 positive control was selected as the known male DNA , and the positive control article in AB - IdentifilerTM kit was selected as the known female DNA ( both concentrations were 0.1 ng / 渭l ) .

The deletion of one allele , deletion of two alleles and deletion of alleles in the low copy DNA typing were performed on the mixed DNA typing , and the likelihood ratio ( LR ) was calculated with reference to the corresponding quantization formula ( 2 ) recommended by Peter Gill .

Results : The results of DNA analysis of 25 cases of mixed samples showed that the number of loci detected in 3 cases was 15 ( detection rate was 100 % ) , the number of 5 cases was 14 ( the detection rate was 93 % ) , the number of 6 cases was 12 ( the detection rate was 73 % ) , the number of the remaining mixed samples was 11 ( the detection rate was 73 % ) .

There was no significant difference in the number of loci , the number of alleles and the number of misjudgment loci .

Of the 17 cases of mixed DNA , the number of loci in 4 cases was 15 , the number of 2 cases was 14 , the number of detection was 13 , the number of 2 cases was 12 , the number of 2 cases was 11 , the lowest detection number was 10 , and 5 cases had no allelic mismatch .

Conclusion : According to the analysis of 44 cases of mixed DNA and 17 cases of mixed DNA , the application range of the restriction combination analysis method is that the relative fluorescence intensity of the highest allele band in 15 loci of mixed DNA typing should be greater than 1000rfu , and the mixed DNA of the male / female two persons with the mixing ratio greater than 30 % . The peak height or peak area information can be analyzed . In the low copy mixed DNA analysis , the quantitative calculation cannot be performed due to the occurrence of the allelic mismatch , and the analysis method is limited in this application .
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：D919

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