产乙酰鸟氨酸脱乙酰基酶基因工程菌的构建及遗传稳定性研究  

Studies on genetic stability of engineering strain producing acetylornithine deacetylase

作　　者：陈悦[1] 李环[1] 韦萍[1] 

机构地区：[1]南京工业大学制药与生命科学学院,江苏南京210009

出　　处：《工业微生物》2008年第2期23-27,共5页Industrial Microbiology

基　　金：国家973项目基金资助(编号2003CB7160004)

摘　　要：利用质粒pET22b(+)为表达载体,成功构建了产N-乙酰鸟氨酸脱乙酰基酶基因工程菌BL21-pET22b(+)-argE,并考察了重组质粒的稳定性。双酶切鉴定了质粒构建正确,SDS-PAGE电泳证实了该菌可高效表达目的蛋白。连续传代50次实验表明重组质粒具有结构稳定性。无选择压力连续传代时,质粒丢失严重;有选择压力时连续传代未发生质粒丢失现象,具有较好的分离稳定性。发酵过程中,用羧苄青霉素代替氨苄青霉素,质粒稳定率由77.78%提高到86.42%。羧苄青霉素浓度为200μg/mL时,质粒稳定率提高到98.33%。Using pET22b（ ＋ ） as the expressing vector, the genetic engineering strain BL21-pEY22b（ ＋ ）-argE produdng acetylomithine deacetylase was successfully constructed, and the genetic stability of the strain was studied. Bienzymatic electrophoretogram showed that the recombinant pla.maid was properly constructed, and SDS-PAGE convinced the strain could overepress the target protein. The recombinant strain was continuously incubated for 50 generations. The results showed the plasmid had structural stability. The plasmid lost seriously without antibiotic, whereas it bad high segregational stability with antibiotic. During fermentaion process, the plasmid stable rate increased from 77.78% to 86.42% when carbenicillin was used instead of ampicillin. The plasmid stable rate was up to 98.33% using 200/μg/mL carbenicillin.

关 键 词：乙酰鸟氨酸脱乙酰基酶 基因工程菌 重组质粒 遗传稳定性 

分 类 号：TQ920.1[轻工技术与工程—发酵工程]