双歧杆菌原生质体的制备及其转化系统的建立  

Prepareation and Transformation of Bifidobacteria Protoplast

作　　者：万翠香[1,2] 章昭琳[1] 王报贵[1] 魏华[1,2] 

机构地区：[1]南昌大学食品科学与技术国家重点实验室,江西南昌330047 [2]南昌大学中德联合研究院,江西南昌330047

出　　处：《食品与发酵工业》2012年第6期61-65,共5页Food and Fermentation Industries

基　　金：国家自然科学基金项目"双歧杆菌丝氨酸蛋白酶抑制剂的功能研究"(30900038);国家"863"计划专题"具有高黏附;拮抗和抗炎症能力的双歧杆菌的高效筛选和功能基因的定位研究"(2008AA10Z337);国家自然科学基金项目"磁珠法和量子点标记筛选及验证双歧杆菌的粘附蛋白"(31000048);2009年江西省主要学科学术和技术带头人培养计划"双歧杆菌Serpin黏附和免疫作用的研究";江西省教育厅科研基金"新型双歧杆菌功能蛋白Serpin的研究"(GJJ10379)

摘　　要：考察了长双歧杆菌WBL001菌体生长状态、酶解浓度、时长、温度以及不同渗透压稳定剂等因素,获得长双歧杆菌WBL001原生质体制备及再生的优化条件:长双歧杆菌以3%接种量活化至MRS培养基,培养至对数中期(OD600∶1),5μg/mL变溶菌素,37℃孵育30 min,以原生质体稳定液SMM为反应缓冲液,原生质体生成率达到81%,其再生率达到48%;在此基础上,通过聚乙二醇PEG融合,将穿梭表达载体pBAX转入双歧杆菌原生质体,成功建立双歧杆菌转化体系。Six factors involved in the preparation and regeneration of Bifidobacteria longum protoplast were investigated,including state of growing cells,Mutanolysin concentration,enzymolysis time,hydrolysis temperature,and osmotic stabilizers.The optimum conditions were as follows： a log phase（19 h） culture of the recipient strain was grown in MRS medium,Mutanolysin concentration was 5 μg/mL,enzymolysis time was 30 min,SMM was used as reaction buffer.Using these optimum conditions,the protoplast formation rate reached 81%,and the ratio of regeneration was 48% in the optimum regeneration medium.On this basis,a Bifidobacteria transformation system of pBAX with PEG was established.

关 键 词：双歧杆菌 原生质体 再生 变溶菌素 转化 

分 类 号：TS201.3[轻工技术与工程—食品科学]