鏈球菌噬菌體裂解酶在大腸桿菌中表達的初步研究

生物工程論文編號:SW078  論文字數:11888,頁數:22

目  錄
目  錄
 中文摘要 I
 英文摘要
 目錄 Ⅲ
 1．緒論 1
 1.1鏈球菌產生疾病的現狀 1
 1.2噬菌體裂解酶簡介 1
 1.3噬菌體裂解酶的作用 2
 1.4本研究的意義 3
 2 實驗部分 4
 2.1實驗材料及儀器 4
器 5
 2.2 實驗方法 5
 3 結果與討論 10
 3.1工程菌的發酵 10
 3.2不同誘導時間對重組菌表達量的影響 12
 3．3 誘導劑用量不同對重組菌表達量的影響 13
 4．總結與展望 14
 致謝 16
 參考文獻 17

摘 要：本文研究重組菌plyC高密度發酵在大腸桿菌中誘導表達的條件。首先利用自控式5L發酵罐進行鏈球菌噬菌體裂解酶A鏈（plyCA）工程菌補料式發酵，對補料時機、誘導時間、誘導劑量對鏈球菌噬菌體裂解酶(plyC)A鏈和B鏈（plyCA,plyCB）在大腸桿菌中表達情況的影響進行研究，以SDS-PAGE檢測方法來檢測目的蛋白表達情況。結果表明，補料時機在發酵進行3.5 h后加入，以一定速度連續加料，并加入誘導劑培養。另以1.7 mmoL/L IPTG誘導表達實驗表明，誘導時間為4小時鏈球菌噬菌體裂解酶重組菌表達的目的產物量最高。用30 µL IPTG誘導使其終濃度為1 mmoL/L可使PLYCA和PLYCB的表達量相對較高。
 關鍵詞：鏈球菌噬菌體裂解酶；大腸桿菌；高密度發酵；誘導表達
 
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.