含增强型绿色荧光蛋白(EGFP)基因的家蚕同源重组质粒载体的构建

Construction of a Homologous Recombination Plasmid Vector pMD-Fib-IE-EGFP Carrying EGFP as Reportor Gene

以含家蚕丝心蛋白重链基因同源片段的质粒pG35 0为出发质粒 ,插入增强型绿色荧光蛋白 (EGFP)基因 ,构建成以EGFP为报告基因的同源重组质粒载体 pMD Fib IE EGFP ,通过PCR及酶切鉴定表明EGFP以正确的方式插入到原始质粒中 ,通过脂质体介导法转染胃癌细胞能发出很强的绿色荧光 ,表明该质粒能够在真核细胞中表达 。

A new plasmid vector pMD-Fib-IE-EGFP which contains enhanced green fluorescent protein gene as reportor gene was constructed by subtitute EGFP for GFP in the original plasmid pG350. The recombinant plasmid was identified by restriction endonuclease enzyme analysis and PCR. The recombined plasmid was transferred to stomach cancer cell mediated by lipofectine. The transfected cell expressed strong fluorescence. The constructed plasmid vector provided foreign DNA material for transgenic silkworm.