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目的 :获得编码日本血吸虫大陆株磷酸丙糖异构酶 ( Sjc TPI)的 c DNA基因片段 ,对其克隆与测序。方法 :以日本血吸虫大陆株成虫总 RNA为模板 ,经过逆转录 -聚合酶链 (式 )反应( RT- PCR)获得编码 Sjc TPI抗原的 c DNA片段 ,并克隆于载体 M13mp18、M13mp19,双脱氧链末端终止法测 DNA序列。结果 :体外 PCR扩增获得了编码 Sjc TPI的 0 .75kb c DNA片段 ,6个重组克隆子经酶切鉴定均有 0 .75kb片段的插入。测得编码 Sjc TPI的 DNA序列。结论 :用自行设计的引物成功地扩增了编码日本血吸虫大陆株 TPI抗原的基因片段并进行了克隆、测序 ,为制备重组 TPI进行疫苗试验打下基础。
OBJECTIVE: To obtain the c DNA gene fragment encoding Sjc TPI of Schistosoma japonicum, and to clone and sequence it. Methods: The cDNA encoding the Sjc TPI antigen was obtained by reverse transcription polymerase chain reaction (RT-PCR) using the total RNA of Schistosoma japonicum adult strains as a template and cloned into vector M13mp18, M13mp19, dideoxy chain Terminal termination assay DNA sequence. Results: The 0. 75kb c DNA fragment encoding Sjc TPI was obtained by in vitro PCR amplification. The 6 recombinant clones were confirmed by restriction enzyme digestion. The DNA sequence encoding the Sjc TPI was determined. Conclusion: The gene fragment encoding the TPI antigen of Schistosoma japonicum mainland strain was successfully amplified by using the designed primers and cloned and sequenced, which laid the foundation for the vaccine preparation of recombinant TPI.