Yoshida K, Sekiguchi K, Mizuno N, Kasai K, Sakai I, Sato H, Seta S
National Research Institute of Police Science, Tokyo, Japan.
Forensic Sci Int. 1995 Mar 21;72(1):25-33. doi: 10.1016/0379-0738(94)01668-u.
This investigation was undertaken as an efficient method for isolating sperm DNA from a mixed fluid sample which contains vaginal epithelial cells in a greater amount. The modified method of the two-step differential extraction procedure was found to be suitable for separating sperm DNA and vaginal epithelial cell DNA from the mixed stains. As the first step of digestion, vaginal epithelial cells in the mixed stains were lysed with Proteinase K and SDS, and sperm heads remaining in the lysed solution were collected by centrifugation. As the second step digestion, the sperm heads were lysed with the buffer containing Proteinase K, SDS and DTT as reducing agent. DNA fractions extracted from the two lysed solutions were enriched, one with sperm DNA and the other with vaginal epithelial cell DNA. MCT118(D1S80), ApoB VNTR and HLADQ alpha types of sperm DNA were detected and were confirmed by matching with corresponding male blood DNA. In the case of vaginal secretion mixed with semen of two males, the mixture of MCT118 types of the two males was detected in sperm DNA fraction.
本研究旨在寻找一种有效的方法，从含有大量阴道上皮细胞的混合液样本中分离精子DNA。结果发现，改良的两步差异提取程序适用于从混合污渍中分离精子DNA和阴道上皮细胞DNA。作为消化的第一步，用蛋白酶K和十二烷基硫酸钠（SDS）裂解混合污渍中的阴道上皮细胞，通过离心收集裂解液中剩余的精子头部。作为第二步消化，用含有蛋白酶K、SDS和作为还原剂的二硫苏糖醇（DTT）的缓冲液裂解精子头部。从两种裂解液中提取的DNA组分得到富集，一种富含精子DNA，另一种富含阴道上皮细胞DNA。检测了精子DNA的MCT118（D1S80）、载脂蛋白B可变数目串联重复序列（ApoB VNTR）和人白细胞抗原DQα（HLADQα）类型，并通过与相应男性血液DNA匹配进行了确认。在阴道分泌物与两名男性精液混合的情况下，在精子DNA组分中检测到两名男性的MCT118类型的混合物。
