Profiler Plus试剂盒扩增Amelogenin基因座变异1例

目的探讨日常检案及DNA数据库建设中常用的ProfilerPlus系统扩增法医生物检材可能出现的变异情况。方法分别采用ProfilerPlus试剂盒扩增、毛细管电泳分析及Amelogenin单基因座扩增、银染技术分型两种方法分别检测两份血痕的基因型。结果应用ProfilerPlus试剂盒扩增、毛细管电泳分析发现X-Y同源Amelogenin基因座的X片段丢失,而运用Amelogenin单基因座扩增、银染技术分型则X片段正常出现。结论应用ProfilerPlus系统分析法医生物检材,有可能出现等位基因丢失现象,此现象需要引起我们在日常检案及DNA数据库建设中的足够重视。     

等位基因特异性PCR技术及其法医学应用

等位基因特异性PCR（allele-specific polymerase chain reaction,AS-PCR）是一种基于等位基因特异性引物引导的PCR技术,可以有效地分析单核苷酸多态性（SNP）,包括碱基的转换、颠换以及插入/缺失多态性,在疾病研究、分子诊断以及法医物证学研究中具有很好的应用价值.本文系统地综述了AS-PCR技术的原理、检测手段、改进方法及在常染色体、Y染色体和线粒体SNP等领域的研究成果,探讨其法医学应用价值.     

判别函数在同胞鉴定中的应用

目的探讨判别函数法在全同胞与半同胞鉴定中的应用价值。方法根据360对全同胞、90对半同胞及360对无关个体的15个STR基因座分型结果,计算全不同(X0)、半相同(X1)和完全相同(X2)的基因座数目,分别根据DFS1=3.898X0+3.973X1-19.481,DHS1=5.687X0+5.300X1-35.112及DR1=7.309X0+5.533X1-44.941的全同胞/半同胞/无关个体判别函数、DFS2=3.872X0+3.931X1-18.895及DR2=7.303X0+5.473X1-44.298全同胞/无关个体判别函数和DHS3=10.227X0+10.436X1-66.102及DR3=11.863X0+11.089X1-79.494半同胞/无关个体判别函数进行判别。结果判别准确率:①用全同胞/半同胞/无关个体判别函数,全同胞组为83.61%,半同胞组为81.11%,无关个体组为83.06%;②用全同胞/无关个体判别函数,全同胞组为96.39%,无关个体组为98.61%;③用半同胞/无关个体判别函数,半同胞组为88.89%,无关个体组为85.00%。结论本文3种判别函数可应用于同胞鉴定,尤其运用全同胞/无关个体判别函数判断同胞关系准确率高,有较高的应用价值。     

Molecular characterisation and population genetics of the DYS458 .2 allelic variant

We recently found a number of intermediate DYS458 alleles, indicated as .2. This allelic variant is distributed in several populations, but currently no information is available regarding the molecular structure and the genealogical correlation of chromosomes with this variant. The molecular characterisation of such allele, its worldwide distribution and the correlated evolutionary history are the subject of the present paper. Molecular and genealogical data are suggestive of a single origin for the .2 variant. Phylogeographic analysis points to either a Middle East or East African origin, but additional data is necessary to clarify this point. Our results suggest that the .2 variants is a stable polymorphism and that it could be used for population studies.     

A D19S433 primer binding site mutation and the frequency in Japanese of the silent allele it causes

Short tandem repeat studies are powerful tools for parentage analysis and for identification of missing persons, victims of murder, and victims of mass fatalities when reference samples are unavailable. The primer in the Identifiler kit failed to amplify an allele at the D19S433 locus, producing a silent ("null") allele. The causal mutation is a base change (G>A) 32 nucleotides downstream from the 3' end of the AAGG repeats. The silent alleles are problematical in parentage analysis because when transmitted, they can cause a parent-child inconsistency that is unrelated to Mendelian genetics. The inconsistency is sometimes termed an "apparent opposite homozygosity" and it produces false evidence of nonparentage. Alternative primers were designed to amplify the D19S433 locus alleles and they detect the silent allele. Frequencies of the (no longer) silent allele were determined to be 0.0114 in 176 people from Shizuoka (Honshu) and 0.0128 in 156 people from Okinawa.     

Allelic alterations of STRs in archival paraffin embedded tissue as DNA source for paternity testing

Owing to a wrong name registered on ID card, the identity of a businessman who had been dead and cremated was suspected, which led his son failed to get legacy. In order to prove the parenthood, the son submitted the gastric cancer tissues surgically removed and embedded in a paraffin block as DNA source for paternity test. After extracting DNA with QIAamp DNA Blood Mini Kit, the 16 STR loci was amplified by two commercial kits of Sinofiler^® (ABI)and Powerplex 16 (Promega), respectively. Both of the STR profiles were similarly showing allelic imbalance pattern at some loci and an additional allele at locus D18S51. The cancerous tissues and adjacent normal tissues were then partitioned off from each other by microscopic analysis of H.E. stained sections and followed by DNA extracting and STR typing, respectively. The allelic alteration could not be found in normal tissues whereas it did in cancerous tissues whose STR profile showed complete loss of one allele (LOH) at loci D13S317 (allele 11 was lost), partial loss of one allele (pLOH) at loci D21S11, D7S820, D19S433, vWA, D12S391 and Amelogein and occurrence of an additional allele (allele 20 was added) at locus D18S51. The results demonstrated that the Paraffin Embedded cancer Tissue used as DNA source for forensic identification is possibly questionable because of their microsatellite instability (MSI) or loss of heterozygosity. It was suggested to partition the normal tissues from the cancer tissues by microscopic evaluation first and then analyzing DNA separately. Comparing the STRs profile of normal tissue with the son's blood sample, the final conclusion was acquired that the donor of the paraffin embedded tissues is the biological father of the son.     

片段长度差异等位基因特异性PCR——一种改良的SNP分型新方法

目的建立一种基于等位基因特异性PCR原理的改良SNP分型新方法:片段长度差异等位基因特异性PCR,并考察特异性引物的3'端第3位、第4位碱基错配对特异性延伸的影响。方法以SNP位点rs759117和rs760887为例,设计两条长度不同、3'末端分别与SNP两个等位基因碱基配对的上游引物,同时在两个等位基因特异性引物3'端第3或第4位碱基引入错配以增加特异性,下游为公用引物。PCR产物经聚丙烯酰胺凝胶电泳、银染显带后确定样本的基因型。结果不同SNP纯合子为长度不同的单一谱带,杂合子则为两条带,其结果与直接测序完全一致。两条特异性上游引物3'端第3或第4位碱基引入错配后非特异性延伸显著减少,且对PCR反应条件的严格性要求明显降低。结论片段长度差异等位基因特异性PCR是一种简单快速而有效的SNP分型新方法;两条特异性引物3'端第3、第4位碱基引入错配可使特异性显著增加     

绝经后妇女肾虚证与载脂蛋白E基因多态性的相关性

目的探讨绝经后妇女肾虚证与载脂蛋白E（apolipoprotein E，ApoE）基因多态性的相关性。方法以80例绝经后肾虚证妇女及65例绝经后无肾虚证妇女为研究对象，以ApoE基因为候选基因，运用限制性片段长度多态性检测ApoE基因型。结果肾虚组人群ApoE基因型ε2/ε3、ε2/ε4、ε3/ε3、ε3/ε4和ε4/ε4频率分别为11．25％、8．75％、58．75％、11．25％、10．00％，对照组人群ApoE基因型ε2/ε3、ε2/ε4、ε3/ε3、ε3/ε4、ε4/ε4频率分别为15．38％、6．15％、69．23％、9．24％、0；肾虚组人群ApoE等位基因ε2、ε3、ε4频率分别为10．00％、70．00％、20．00％，对照组ε2、ε3、ε4频率分别为10．76％、81．55％、7．69％；肾虚组ApoE等位基因ε4频率明显高于正常对照组（χ^2=4．386，P〈0．05）。结论ApoE等位基因ε4频率升高与绝经后妇女肾虚证相关，可能是肾虚证的危险因子；ApoE基因型ε4／ε4是绝经后妇女肾虚证一个有用的标志物。