小鼠内皮抑素基因克隆、测序及pEgr-IFNγ-mEndostatin重组双基因表达质粒的构建

Cloning and sequencing of mouse endostatin and construction of recombinant plasmid pEgr-IFNγ-mEndostatin

目的 :克隆小鼠内皮抑素 (m Endostatin)编码区 c DNA序列并构建含 Egr- 1启动子的 IFNγ和 m Endo-statin双基因表达载体。方法 :利用逆转录多聚酶链反应法 (RT- PCR) ,以小鼠肝细胞 m RNA为模板 ,扩增获得全长 m Endostatin,与 p MD1 8T载体连接作全自动测序 ,并利用基因重组技术构建含 Egr- 1启动子的 IFNγ和m Endostatin双基因表达质粒。结果 :经测序证实获得的 m Endostatin序列与文献报道完全一致 ,并构建了含 Egr-1启动子的 IFNγ和 m Endostatin双基因表达质粒 p Egr- IFNγ- m Endostatin。结论 :利用 RT- PCR法成功克隆了m Endostatin的 c DNA序列 ,构建了 p Egr- IFNγ- m Endostatin重组双基因表达质粒。

Objective To clone the sequence of the cDNA of mouse endostatin (mEndostatin) coding area and construct an expression vector containing Egr 1 promoter, IFNγ and endostatin genes. Methods Full length mEndostatin was obtained with the technique of RT PCR and using mouse hepatocyte mRNA as template, pMD18T mEndostatin was sequenced automatically and an expression vector containing Egr 1 promoter, IFNγ, and endostatin genes was constructed with gene recombinant technique. Results It was proved that the cloned mEndostatin cDNA to be completely identical with that reported in the literature and the recombinant plasmid containing Egr 1 promoter, IFNγ, and endostatin genes was constructed successfully. Conclusion mEndostatin cDNA was cloned and the expression vector pEgr IFNγ mEndostatin was constructed successfully.