摘要
研究构建人基质金属蛋白酶7(MMP-7)的原核表达载体,并进行原核表达获得重组蛋白MMP-7。以人肾肿瘤组织总RNA为模板,PCR方法获得MMP-7成熟蛋白的基因序列,构建含10xhis标签的原核表达载体,转化大肠杆菌BL21(DE3),并经异丙基-β-D-硫代吡喃半乳糖苷(IPTG)诱导表达,获得重组蛋白,并经十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE),检测其表达情况。最终得到pET24a(+)-MMP-7-10his重组质粒,诱导表达后重组蛋白占总蛋白的41%左右,获得21.2kD大小蛋白,与目的蛋白大小一致,纯度大于90%,为进一步研究奠定基础。
To obtain the recombinant matrix metalloproteinase 7 (MMP-7), this article using the prokaryotic expression and constructing the prokaryotic expression vector. The total RNA of human kidney tumor tissue was used as a template for reverse transcription, after PCR amplification, the target fragment was digested and cloned into the prokaryotic expression vector pET24a( + ) to construct the recombinant plasmid pET24a( + )-MMP-7- 10his,then the recombinant plasmid was transferred into BL21(DE3) and the MMP-7 expression was induced by IPTG and examined by SDS-PAGE. We obtained the recombinant plasmid of human MMP-7 gene. The expressed MMP-7 recombinant protein accounted for approximately 41% of the total bacteria proteins, and the purity of MMP-7 was above 90%.
出处
《安徽农学通报》
2013年第7期31-33,107,共4页
Anhui Agricultural Science Bulletin