鸭疫里默氏杆菌和大肠杆菌双型融合菌株的构建  

Establishment of protoplast fusion cells of Riemerella anatipestifer and Esherichia coli

作　　者：王春平[1,2,3] 韦强[1] 鲍国连[1] 崔言顺[2] 刘燕[1] 邵泽香[1,2] 季权安[1] 肖琛闻[1] 李建亮[2] 

机构地区：[1]浙江省农业科学院畜牧兽医研究所,浙江杭州310021 [2]山东农业大学动物科技学院,山东泰安271018 [3]山东日照市畜牧兽医局,山东日照276826

出　　处：《中国兽医学报》2010年第7期903-907,共5页Chinese Journal of Veterinary Science

基　　金：浙江省重点科技攻关资助项目(2006C12026);浙江省农业科学院创新能力提升工程资助项目

摘　　要：以耐药标记的鸭疫里默氏杆菌TXRA1(Chlr,Erys)和大肠杆菌ZBO78(Eryr,Chls)作为亲本菌株,在DnaseⅠ始终存在的条件下,采用Lysozyme-EDTA法制备2菌株的原生质体,原生质体制备率分别达到了92.00%和91.94%,再生率分别达到了38.77%和37.29%。通过PEG法进行2菌株原生质体融合,成功获得31株具有双亲本耐药性(Chlr,Eryr)的融合菌株,并应用双重PCR方法对融合株的部分外膜蛋白A(ompA)基因进行扩增,其中有11株融合菌株能够表达2亲本的ompA基因,有20株仅表达亲本大肠杆菌的ompA基因。融合菌株的形态、染色特性和生化特性等介于2亲本菌株之间,连续传15代后融合菌株的上述性状仍能够稳定遗传。Using Riemerella anatipestifer TXRA1（Chlr,Erys） and Esherichia coli ZBO78（Eryr,Chls） strains as parental strains,protoplasts were prepared with lysozyme and EDTA while Dnase Ⅰ existed,the formation frequency of two parental strains was 91.94% and 92.00% and the regeneration frequency was 37.29% and 38.77%,respectively.PEG method was used for protoplast fusion and 31 fusion cells were obtained with the character of two antibiotics resistance （Eryr,Chlr）.Duplex PCR method was used to detect part of the outer membrane protain A （ompA） gene in the fusion cells,11 fusions had the ompA gene of their two parental strains,another 20 only had the ompA gene of Esherichia coli ZBO78.All fusions had hereditary properties of the two parental strains,such as morphology,staining character,biochemical activities,etc,the hereditary properties of the 31 fusions were found to be stable after 15 times of passages.

关 键 词：鸭疫里默氏杆菌 大肠杆菌 原生质体融合 

分 类 号：S852.61[农业科学—基础兽医学]