论文部分内容阅读
目的:在前期克隆丹参SmJAZ1基因的cDNA全长基础上,为研究丹参SmJAZ蛋白的功能,在大肠杆菌BL21(DE3)中诱导表达丹参SmJAZ1蛋白,并对其表达条件进行优化。方法:利用分子克隆的方法将丹参JAZ1基因构建到原核表达载体pET32a上,转化到大肠杆菌BL21(DE3)宿主菌中进行诱导表达。结果:对影响重组蛋白表达的4个因素,即诱导温度、诱导时间、IPTG浓度、及IPTG添加时间进行优化,确定丹参SmJAZ1重组蛋白最适表达条件。IPTG浓度对目的蛋白的表达量没有显著影响;随诱导时间和诱导温度增加,SmJAZ蛋白的表达量增加;而IPTG添加时间对蛋白的表达量有明显影响。结论:丹参JAZ1蛋白在30℃温度条件下,重组菌生长2 h(A600=0.9),加入0.1 mmol.L-1浓度的IPTG,诱导20 h后,表达条件最合适。
OBJECTIVE: To study the function of SmJAZ1 protein in Salvia miltiorrhiza Bunge, the expression of SmJAZ1 protein was induced in E. coli BL21 (DE3) and its expression conditions were optimized. Methods: The Salvia miltiorrhiza JAZ1 gene was cloned into the prokaryotic expression vector pET32a by molecular cloning and transformed into E. coli BL21 (DE3) host strain for induction. Results: Four factors affecting the expression of recombinant protein, namely induction temperature, induction time, IPTG concentration and IPTG addition time, were optimized to determine the optimal expression conditions of SmJAZ1 recombinant protein. The concentration of IPTG had no significant effect on the expression of the target protein. The expression of SmJAZ protein increased with the induction time and the induction temperature, while the IPTG addition time had a significant effect on the protein expression. CONCLUSION: Recombinant bacterium grows for 2 h (A600 = 0.9) at 30 ℃, and the optimal conditions for the expression of Salvia miltiorrhiza JAZ1 protein after adding 0.1 mmol·L-1 IPTG for 20 h.