鸡生长激素受体胞外区原核表达分析

Expression Analysis of Chicken Growth Hormone Receptor Extracellular Region in Protokaryon

生长激素与其靶细胞膜表面的生长激素受体(GHR)结合,可以促进动物生长发育和调节新陈代谢。为获得鸡GHR蛋白,试验通过PCR扩增获得GHR胞外区序列。经双酶切后连接入原核p ET-32a(+)表达载体,构建了鸡GHR胞外结构域的原核表达载体。将该重组质粒转入E. coli BL21(ED3)表达菌株。通过IPTG诱导其表达GHR重组蛋白,并探讨不同IPTG浓度、诱导时间和诱导温度等条件对鸡GHR重组蛋白表达的影响。结果表明:诱导GHR重组蛋白表达的最适条件为IPTG终浓度2.00 mmol/L、诱导时间为4 h、培养温度为37℃,在此条件下,GHR重组蛋白表达量最高。研究结果为后续GHR抗体制备奠定了基础。

Growth hormone(GH) can promote animal growth and regulate metabolism by binding with the growth hormone receptor(GHR) on the membranes of the target cell. In order to obtain chicken GHR protein, we firstly conducted PCR amplification to get the extracellular domain of GHR. After digesting with double enzymes, the PCR product was linked to pET-32 a(+) vector. Then, the recombinant plasmids were transferred into E. coli BL21(ED3).Subsequently, the GHR recombinant protein was inducted for expression by adding IPTG. Effects of IPTG concentration,induction time and temperature on the expression of GHR recombinant protein were studied. The results showed that the optimum conditions for the expression was IPTG with a final concentration of 2.00 mmol/L, induced at 37 ℃ for 4 h.This study laid a basis for producing chicken GHR antibody.