利用SOE-PCR与TD-PCR技术对气肿疽梭菌FliA(C)-NanA融合基因扩增方法的构建  

A Method Construction of Amplification Fusion Gene FliA(C)-NanA of Clostridium chauvoei by SOE-PCR and TD-PCR Technology

作　　者：李香春[1] 金鑫[1] 朴春宇 金成德 朴春实 

机构地区：[1]延边大学农学院动物医学系,吉林延吉133000 [2]延吉市动物卫生检疫站,吉林延吉133000 [3]吉林省晖春畜牧管理局,吉林晖春133000 [4]吉林省珲春市动物卫生监督所,吉林晖春133300

出　　处：《安徽农业科学》2013年第24期9921-9923,共3页Journal of Anhui Agricultural Sciences

摘　　要：[目的]构建联合表达气肿疽梭菌的鞭毛蛋白与分泌蛋白神经氨酸酶的方法,用以制备气肿疽生物工程亚单位疫苗。[方法]根据已公布的气肿疽梭菌FliA(C)和NanA序列设计引物,联合使用SOE-PCR与TD-PCR技术,建立融合基因FliA(C)-NanA的扩增方法。[结果]试验成功扩增出了融合基因FliA(C)-NanA。[结论]SOE-PCR方法无需在引物设计中加入酶切位点进行连接,降低了试验成本,且2个基因间未加入其他核酸序列,避免了表达出影响免疫效果的蛋白;TD-PCR的应用节省了试验时间,避免了筛选退火温度的繁琐步骤,解决了基因丢失的问题。[ Objective] In order to prepare gene engineering vaccines of blackleg, a method to combined express flagellin and secretory protein neuraminidase of Clostridium chauvoei was constructed. [ Method ] According to the published FliA （C） and NanA sequences of C. chauvoei, the primers were designed. Then using SOE-PCR combined with TD-PCR technology, the amplified method of fusion gene FliA（ C ）-NanA was con- structed. [ Result ] The fusion gene FliA （ C ） -NartA was amplified successfully. [ Conclusion ] The SOE-PCR method without restriction sites to connect in the primer design, reduced the cost of the experiment, and between the two genes did not join the other nucleic acid sequences, to avoid the expression of protein 9.fleeted immune effect; the application of TD-PCR saved the test time, and could avoid tedious steps for screening of an- nealing temperature and the gene loss problem.

关 键 词：气肿疽梭菌 融合基因 SOE-PCR TD-PCR 

分 类 号：S188[农业科学—农业基础科学]