链球菌噬菌体裂解酶在大肠杆菌中表达的初步研究

论文编号:SW078论文字数:11888,页数:22

摘 要：本文研究重组菌plyC高密度发酵在大肠杆菌中诱导表达的条件。首先利用自控式5L发酵罐进行链球菌噬菌体裂解酶A链（plyCA）工程菌补料式发酵，对补料时机、诱导时间、诱导剂量对链球菌噬菌体裂解酶(plyC)A链和B链（plyCA,plyCB）在大肠杆菌中表达情况的影响进行研究，以SDS-PAGE检测方法来检测目的蛋白表达情况。结果表明，补料时机在发酵进行3.5 h后加入，以一定速度连续加料，并加入诱导剂培养。另以1.7 mmoL/L IPTG诱导表达实验表明，诱导时间为4小时链球菌噬菌体裂解酶重组菌表达的目的产物量最高。用30 µL IPTG诱导使其终浓度为1 mmoL/L可使PLYCA和PLYCB的表达量相对较高。

关键词：链球菌噬菌体裂解酶；大肠杆菌；高密度发酵；诱导表达

Primary Research of Streptococcal Bacteriophage Lysin in Escherichia coi

Abstract：This article to investigate the optimal high-density fermentation of recombinant bacteria plyC induced in E. coli conditions. Firstly, make use of 5L autocontrol fermentor for streptococcal bacteriophage lysin chain A (plyCA) fed batch fermentation bacteria feeding on feeding time, induced time, induced dose of the streptococcus bacteriophage lyase (plyC) A chain and B chain ( plyCA, plyCB) expressed in E. coli to study the impact of the situation to SDS-PAGE detection method to detect protein expression. The results show that the feeding time after 3.5 h in the fermentation by adding, to a certain speed continuous feeding, and the addition of induction agent training. The other to 1.7 mmol/L IPTG induced expression experiments show that the induction time of 4 hours lyase of streptococcal bacteriophage lysin recombinant expression produced of the highest purpose. Induced with 30 μL IPTG to a final concentration of 1 mmoL/L may cause PlyCA and the PlyCB expression quantity is relatively high

Key words: PLYC; streptococcal bacteriophage lysin;. E.coil; high cell-density fermentation; Induction expression.

目 录

目 录

中文摘要 I

英文摘要

目录 Ⅲ

1．绪论 1

1.1链球菌产生疾病的现状 1

1.2噬菌体裂解酶简介 1

1.3噬菌体裂解酶的作用 2

1.4本研究的意义 3

2 实验部分 4

2.1实验材料及仪器 4

器 5

2.2 实验方法 5

3 结果与讨论 10

3.1工程菌的发酵 10

3.2不同诱导时间对重组菌表达量的影响 12

3．3 诱导剂用量不同对重组菌表达量的影响 13

4．总结与展望 14

致谢 16

参考文献 17