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人可溶性白介素-1受体真核表达载体pcDNA3/sIL-1R的构建 被引量:4
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作者 李鹏 徐艳 +2 位作者 张蕴惠 章锦才 王大章 《华西口腔医学杂志》 CAS CSCD 北大核心 2003年第2期140-143,共4页
目的 本研究旨在构建人可溶性白介素_1受体 (sIL_1R)真核表达载体pcDNA3 sIL_1R ,为研究骨关节病的基因治疗奠定实验基础。方法 将人sIL_1R基因的RT_PCR纯化产物及质粒pcDNA3 1(+)DNA经KpnI和XhoI双酶切、胶回收纯化酶切片段后 ,体外... 目的 本研究旨在构建人可溶性白介素_1受体 (sIL_1R)真核表达载体pcDNA3 sIL_1R ,为研究骨关节病的基因治疗奠定实验基础。方法 将人sIL_1R基因的RT_PCR纯化产物及质粒pcDNA3 1(+)DNA经KpnI和XhoI双酶切、胶回收纯化酶切片段后 ,体外连接酶切片段 ,使其定向重组 ,再将重组DNA转化E .coliCompetentCellsJM10 9,经复苏后 ,在含Ampr 的LB固体培养基上筛选出阳性克隆。结果 挑取的LB固体培养基上的 6个单菌落经酶切及测序鉴定 ,证实均为阳性克隆 ,即sIL_1R与pcDNA3 1(+)体外重组成功。结论 采用体外重组技术 ,成功地将人sIL_1RcDNA插入了真核表达载体pcDNA3 1(+) 展开更多
关键词 人可溶性白介素-1受体 真核表达载体 PCDNA3 骨关节病 基因治疗
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生长激素信号肽可诱导重组蛋白外分泌表达 被引量:4
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作者 张志谦 李金萍 胡颖 《中国生物化学与分子生物学报》 CAS CSCD 北大核心 2005年第2期282-286,共5页
Signal peptide capable of efficiently directing many protein secretion in mammalian cells is one of the key elements in recombinant protein production,gene therapy and the development of DNA vaccines.In order to explo... Signal peptide capable of efficiently directing many protein secretion in mammalian cells is one of the key elements in recombinant protein production,gene therapy and the development of DNA vaccines.In order to explore the possibility of rat growth hormone signal peptide as such an element,a new vector based on the mammalian expression vector pcDNA3 was constructed by employing rat growth hormone(rGH) signal peptide as leading sequence,followed by multiple cloning sites,the myc epitope-tag and 6×his purification tag in the expression cassette.The vector was validated by successfully expressing and secretion of chick MMP-2 C-terminal PEX domain,a potential angiogenesis inhibitor,and tandem peptide repeats of myc epitope-tag in COS-7 cells.These results suggest that rat growth hormone signal peptide is effective in the mediation of recombinant protein expression and secretion,and this vector provides a new tool for universal cloning and secretion of exogenous proteins in mammalian cells. 展开更多
关键词 expression 分泌表达 重组蛋白 生长激素 vector 信号肽 SIGNAL universal elements multiple 诱导 SIGNAL PCDNA3 domain the and MMP-2 COS-7 rat tool new was one key his tag The PEX for ce
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