摘要
目的 :为发展多特异性避孕疫苗 ,制备重组抗原pZP3α -hCGβCTP10 9～ 14 5,用基因工程技术构建重组质粒pPIC9K -pZP3α -hCGβCTP10 9～ 14 5.方法 :根据pZP3α全长和hCGβCTP10 9～ 14 5编码的DNA列设计两对引物 ,通过部分重叠聚合酶链式反应 (overlappingPCR) ,扩增出pZP3α -hCGβ -CTP10 9～ 14 5DNA片断 ,克隆到载体质粒pPIC9K中 ,然后导入大肠杆菌DH5α ,用PCR和双酶切反应鉴定重组质粒 ,并用DNA测序仪对重组质粒测序 .结果 :3步PCR扩增出pZP3α -hCGβ -CTP10 9～ 14 5DNA片段 ,插入到载体质粒pPIC9K的克隆位点 ,获得重组pPIC9K -pZP3α -hCGβ -CTP10 9～ 14 5表达质粒 ,测序结果显示插入序列与设计预期完全一致 .结论 :重组质粒pPIC9K -pZP3α -hCGβ -CTP10 9～ 14 5构建成功 ,为表达重组抗原pZP3α -hCGβ -CTP10 9～ 14 5。
Aim:Prepare antigen pZP3α-hCGβ-CTP 109～145 for developing contraceptive vaccine which reacts with zona pellucida and hCG to construct recombinant plasmid pPIC9K-pZP3α hCGβ-CTP 109～145 by DNA recombination. Methods: Two pairs of primers (P1/P2,P3/P4) were synthesized basing on pZP3α and hCGβ-CTP 109～145 cDNA sequences. The pZP3α-hCGβ-CTP 109～145 chimera DNA fragment was prepared by overlapping PCR. The chimera DNA fragment was inserted into pPIC9K plasmid and transformed into E. coli DH5α. The recombinant plasmid pPIC9K-pZP3α hCGβ-CTP 109～145 was identified for its inserted DNA sequence and orientation by PCR, EcoRI and NotI digestion, and DNA sequencing. Results: pZP3α-hCGβ-CTP 109～145 DNA fragment was synthesized by three steps of overlapping PCR and was inserted into vector plasmid pPIC9K. The result of the sequencing was entirely consistent with anticipation. Conclusion:Recombinant plasmid pPIC9K-pZP3α hCGβ-CTP 109～145 was constructed successfully.
出处
《暨南大学学报（自然科学与医学版）》
CAS
CSCD
2002年第3期100-106,共7页
Journal of Jinan University(Natural Science & Medicine Edition)
基金
国家"九.五"计划生育科技攻关项目 (96 - 90 4 - 0 1- 0 1)
