中国寄生虫学与寄生虫病杂志 ›› 1990, Vol. 8 ›› Issue (4): 264-266.

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用特异性抗体筛选日本血吸虫基因组DNA基因库

陶伊文,徐裕信,刘述先,郑兆鑫,严维耀   

  1. 中国预防医学科学院寄生虫病研究所; 中国预防医学科学院寄生虫病研究所; 中国预防医学科学院寄生虫病研究所; 复旦大学遗传学研究所; 复旦大学遗传学研究所
  • 出版日期:1990-11-30 发布日期:2017-01-09
  • 基金资助:
    国家科委863高技术计划资助项目

SCREENING OF SCHISTOSOMA JAPONICUM ADULT WORM GENOMIC DNA LIBRARY BY SPECIFIC ANTIBODY

  • Online:1990-11-30 Published:2017-01-09

摘要: 用酶免疫测定法筛选日本血吸虫(S.j.)基因组DNA基因库,先将λ噬菌体(λgt11),重组体噬菌斑的表达抗原转移至硝酸纤维(NC)膜,然后用经导入λgt11的大肠杆菌Y1090细胞裂解液吸收的感染兔血清(IRS)检测。结果,从97只平板初筛出8个可能阳性噬菌斑,经再次筛选,8个中2个仍为阳性。其中1个阳性克隆经纯化后导入Y1089细胞扩增,其表达产物经ELISA复筛进一步证实S.j.抗原阳性。表明该克隆编码S.j.抗原,该克隆的表达产物能被IRS识别。该免疫筛选方法能有效地分离单个编码S.j.抗原的克隆,且筛选效率高(每次可筛选1×10~5噬菌斑形成单位),结果可靠、特异,不需要同位素标记技

关键词: 日本血吸虫, 基因组DNA基因库, 基因库筛选, 抗体探针

Abstract: S. japonicum adult worm genomic DNA libraries were screened by enzymeimmunoassay. The antigens produced by recombinant λgt 11 plaques were transferred to nitrocellulose filters. Clones encoding given antigens were detected with infected rabbit sera (IKS) which were preabsorbed with lysate of induced λgt 11 in Y1090 cells. Eight putative positives were picked up from 97 plates in the primary screening and were rescreened a second time, and, 2 of them consistently gave good positive signals in subsequent screenings. One of the phage clones was purified to homogeneity and mixed with Y1089 cells. The expressed products still showed positive when rescreened by ELISA, indicating that the clone encor ding S. Japonicum antigen could be recognized by IRS. The immmunoscreening method described here was able to efficiently isolate single clones encoding S. japonicum antigens The method has the advantages of screening libraries efficiently, reliably and specifically, and it might open the way to screen genes encoding S. japonicum antigens inducing protective immunity (Fig. 1).