【摘要】 目的：从日本血吸虫虫卵文库中筛选并鉴定出与免疫诊断及免疫预防有关的基因克隆。方法：采用Ｓ．ｊＳＥＡ超免疫兔血清对日本血吸虫卵文库进行筛选。先去除抗大肠杆菌抗体，经三轮筛选得到１２个阳性克隆，然后，用辅助噬菌体进行体内剪切，经抗菌素平板筛选含重组质粒的阳性菌落，每个菌落分为２份，一份进行ＰＣＲ扩增以测定插入片段的大小。另一份用于制备纯质粒ＤＮＡ模板，进行ＤＮＡ序列测定，通过ＧＣＧ软件对所得ＤＮＡ序列与ＧｅｎＢａｎｋ中的序列进行同源性比较。结果：共筛得１２个阳性克隆，ＰＣＲ后测得其长度大多为１．２ｋｂ，其中８个与Ｓ．ｊ热休克蛋白７０的基因同源，２个与Ｓ．ｊ钙网蛋白同源，１个与Ｓ．ｍｉｍｍｕｎｏｐｈｉｌｉｎ同源，１个与Ｓ．ｊ２３ｋＤａ蛋白同源。结论：本研究首次报道用抗ＳＥＡ血清对日本血吸虫虫卵文库的筛选结果，所得的１２个阳性克隆均为编码日本血吸虫免疫诊断及预防有关的基因。更多还原

【Abstract】 AIM: To screen and identify genes encoding vaccine candidates and molecules with diagnostic potential from S.japonicum(S.j) egg cDNA library. METHODS: S.j egg cDNA library was screened by sera from SEA immunized rabbits. Positive clones were identified after three rounds of screening and were excised automatically in vivo with helper phage. Products of PCR were used to detect insert sizes by agarose gel electrophoresis. DNA samples were obtained by purification using mini prep kit. Automatic sequencing was carried out and the sequence data were analysed using GCG software. RESULTS: Twelve clones were identified as positive and the inserts were around 1.2 kb in size. Compared with DNA sequences in GenBank, eight clones were homologous to a 70 kDa S.j heatshock protein( S.j HsP70). C18 exhibited 97.8% homology with S.j 23. C22 and C23 shared 82.4% and 89.5% homology with S.j calreticulin,respectively. C12 was homologous to S.mansoni(S.m.) immunophilin. CONCLUSION: This paper first reported the results of immunoscreening S.j egg cDNA library by anti SEA sera.Twelve positive clones were found to be related to protective immunity and/or immunodiagnosis of schistosomiasis.更多还原