优化的复合自动诱导培养基（CAI-4）对外源蛋白在大肠杆菌中 表达的影响

Culture optimization for protein expression in Escherichia coli

摘要:

[目的]原核表达系统是目前最为广泛使用的一种外源蛋白表达系统。在利用原核表达系统表达目的蛋白的过程中，可溶性外源蛋白的产量是决定成本和效率的决定性因素。[方法]本项研究中利用本实验室构建的7种不同的重组质粒(-1、p-2、p-3、p-4、p-5、p-6、p-7)，检测其在复合自动诱导培养基中的表达情况，评价哪一种培养基更适合于外源蛋白的表达，提高目的蛋白的产量。[结果]结果显示，这7种重组蛋白在复合自动诱导培养基的表达量是普通LB培养基的4~8倍；并在此基础上，对复合培养基的成份进行进一步优化，形成了一种优化培养基(改良培养基-4)，P-1、P-2、P-3这3种融合蛋白在这种改良培养基中的表达量比优化前提高了至少2倍。

Abstract:

[Objective] Escherichia coli is the best characterized system in every aspect, and it has been the most widely used host for the production of recombinant proteins. Furthermore, the high cell density culture (HCDC) has allowed production of various proteins with high yield and high productivities. Automation and miniaturization are key issues of high-throughput research projects in the post-genomic era. [Methods] We applied a new culture-complex auto-inducing media (CAI) for heterogenous protein expression in E. coli. The CAI is in accord with automation and miniaturization. To test expression efficiency in the CAI, we constructed seven different plasmids named p-1、p-2、p-3、p-4、p-5、p-6 and p-7. These plasmids were transformed into E. coli BL21 (DE3), then expressed in both LB and CAI. [Results] The expression levels of seven fusion proteins in CAI were four times greater than those in LB. To improve the expression level even more, we analyzed the composition of the CAI and optimized the culture. Through a series of changes we formed a new optimized culture (CAI-4). Comparing the expression levels of there fusion proteins (P-1、P-2、P-3) in CA, the expression levels of fusion proteins in CAI-4 were twice greater than the control.