【摘要】 选择一段与COL1A1基因相应序列同源、适合于在大肠杆菌E.coli中表达、含有编码促细胞粘附位点GER三肽密码子的基因序列。利用RT-PCR技术从人组织扩增该序列,构建pET28a(+)-COL重组表达质粒并转化到大肠杆菌BL21(DE3)pLysS。IPTG诱导表达获得重组人源性胶原RHDC,表达量达到菌体总蛋白的40%左右,SDS-PAGE和Western blot结果推测该胶原具有三螺旋结构,细胞粘附试验证明RHDC具有促细胞粘附能力,有应用于生物材料领域的潜力。更多还原

【Abstract】 The DNA sequences, which originated from gene COL1A1, optimized for E.coli codon usage and contained cell adhesion domains of GER units, was amplified by RT-PCR technology from human tissue. The recombinant expression vector pET28a(+)-COL encoded the recombinant human-derived collagen (RHDC) was constructed and transformed host E.coli strain BL21(DE3)pLysS. The RHDC production was induced by the addition of IPTG and approximately 40% of the total strain proteins. RHDC could form a triple-helical structure suggested by the SDS-PAGE and Western blot analysis. Cell adhesion assay indicated that RHDC had high cell adhesive character, which makes it suitable for use as biomaterials.更多还原