摘要
为深入开展人巨细胞病毒（ＨＣＭＶ）感染的分子流行病学和发病机理研究，作者从不同人群获得１０株ＨＣＭＶ，经培养、传代和增殖后提取病毒ＤＮＡ，而后分别用限制性内切酶ＨｉｎｄⅢ和ＥｃｏＲＩ消化，用３２Ｐ标记的ＨＣＭＶＤＮＡＨｉｎｄⅢ亚克隆片段（ｐＣＭ１０１５、ｐＣＭ１０３５）做探针，进行Ｓｏｕｔｈｅｒｎ杂交和限制性片段长度多态性分析。结果显示：该１０株ＨＣＭＶ的主要限制性片段与标准株ＡＤ１６９相同，证明该病毒基因组的结构相当稳定，但其Ｌ－Ｓ结合区和末端片段有变异，表现为酶切位点的丢失或获得；流行病学上相关毒株限制性酶谱相似，而无相关毒株的限制性酶谱则有所不同。由此表明，以ＥｃｏＲＩ酶切片段与ｐＣＭ１０３５探针杂交易于发现其基因组变异。临床毒株的Ｓｏｕｔｈｅｒｎ杂交分析可用于ＨＣＭＶ感染的分子流行病学和发病机理的研究。
Ten clinical isolated strains of Human cytomegalovirus(HCMV) were obtained from 73 urine specimens of different people.Isolations from urine were carried out in human embryolung fibroblasts. Viral isolates were passaged four times.HCMV DNAs of laboratory strain AD 169 and 10 clinical isolated strains were extracted with Hirt method,digested with each of the restriction enzymes EcoRI,HindⅢ.Comparison of restriction fragment length polymorphism(RFLP) of AD 169 and isolated strains were made by hybridizing digested DNA with 32 P labeled with HCMV HindⅢ cloned subgenomic fragments as the probe (pCM1035,pCM1015). pCM1035 is located in the joining region between the long(L) and short (S) unique sequences of the virus (L S junction );pCM1015 is located in the terminal sequences of the virus.The results showed the genomic high degree of homology existed among all strains and the variable restriction site was in the L S junction and terminal portion.The RFLP patterns of the clinical isolates which did not have relation in epidemiology were different,but the patterns of clinical isolates related in epidemiology were quite similar.Polymorphism frequently occurred in this case of EcoRI digested fragment hybridized with the probe of pCM1035.Southern hybridization of HCMV isolations is useful to resarches into the molecular epidemiology and pathogenesis of HCMV infection.
出处
《华西医科大学学报》
CSCD
1996年第2期177-181,共5页
Journal of West China University of Medical Sciences
关键词
人巨细胞病毒
RFLP
DNA
Human cytomegalovirus Restriction fragment length polymorphism Southern hybridization
