骨形态发生蛋白在大肠杆菌中表达的研究

Recombinant human BMP expressed in escherichiacoli

目的骨形态发生蛋白(bone morphogenetic protein,BMP)是一种具有诱导活性的酸性糖蛋白,在组织工程研究和临床应用上需求量较大,BMP-2在组织中含量较微,利用基因工程方法生产重组BMP-2显得尤为重要。方法本研究根据人骨形态发生蛋白(bone morphogenetic protein,BMP)基因序列设计出引物,以克隆载体质粒pBR322-BMP-2为模板,得到BMP-2基因并重组入原核表达载体pGEX-4T-1中,将经酶切和测序鉴定正确的重组质粒转化大肠杆菌DH5α,用IPTG诱导表达,表达产物经SDS-PAGE鉴定并用Western bloting检测。结果SDS-PAGE显示,得到40 KD的目的蛋白,表达量可达菌体总蛋白的15.2%,Western bloting检测表明,抗BMP-2单抗可与相对分子量40 KD大小的电泳条带发生特异性反应,从而证明表达产物具有目的蛋白构型。结论本研究为进一步研究利用原核表达系统大量生产BMP-2打下基础。

Aim The bone mopogenetic proteins （BMPs） area family of growth factors that regulate the development of bone. A large number of BMP-2 needed for both bone tissue engineering ressearch. Thus, the effective genetic engineering way is necessary, produce sufficient BMP-2 protein. Methods To clone BMP-2 gene from the pBR322-BMP-2 for the construction of prokaryotic expression vector pGEX- BMP-2. The recombinant plasmid was transformed into DH5α and IPTG was used to induce the expression of BMP-2. The recombinant protein（BMP-2） was confirmed by SDS-PAGE analysis,western bloting assay. Result The result indicated that the recombinant protein showed expression which was over 15.2% of total protein according to SDS-PAGE, and had configuration of nature BMP-2 with western bloting assay. Conclusion Our research laid a good foundation for nass-producing BMP-2.