【摘要】 目的 构建酵母双杂交用NCLs致病相关基因CLN8的诱饵质粒，并检测其自激活作用。方法 PCR扩增目的基因片段，酶切后与诱饵载体pLexA连接形成重组诱饵质粒，化学转化法将重组诱饵质粒转化到酵母细胞EGY48中，β-半乳糖苷酶滤膜分析检测重组诱饵质粒对报告基因LacZ的自激活情况。结果 扩增出目的基因片段，成功构建了重组诱饵质粒plexA-cln8，经酶切和测序鉴定正确;重组诱饵质粒无自发激活报告基因功能。结论 重组诱饵质粒构建成功，对报告基因无自激活作用，为下一步cDNA文库筛选奠定了基础。

    关键词 CLN8 酵母双杂交 诱饵质粒 自激活检测

    Construction of CLN8bait plasmid and detection of its self-activating

    effect in yeast two hybrid system

    Li Jie，He Shuya，Xiao Weichun，et al.

    Department of Biochemistry and Molecular Biology Rearch，Nanhua University，Hengyang421001.

    【Abstract】 Objective To construct the bait plasmid of CLN8gene in yeast two hybrid system and examine whether the bait plasmid of CLN8has self-activating effect.Methods The fragments of CLN8gene was amplified by PCR.The amplified fragment of CLN8was ligased with the vector pLexA to construct recombinant plasmid plexA-cln8.The recombinant plasmid was transfected into yeast strain EGY48by chemistrytransfection protocol ......