CAJ | 학술논문

目的:构建组蛋白 H2A、H2B、H3、H4的酵母双杂交诱饵载体,检测其在酵母细胞中的表达和自激活活性,为应用酵母双杂交系统筛选与组蛋白相互作用的蛋白奠定基础.方法:扩增 H2A、H2B、H3、H4蛋白基编码区 cDNA序列,克隆至酵母表达载体 pGBK-T7中,再将其转化至酵母细胞 AH109中,提取酵母总蛋白,检测各种组蛋白的表达,并检测表达的组蛋白在酵母细胞中对报告基有无自激活作用.结果:将 H2A、H2B、H3、H4基的 cDNA 克隆至酵母表达载体 pGBK-T7中,其中组蛋白 H4得到正确表达,且对报告基 LacZ、HIS3、Ade 无激活作用.结论:可以利用酵母双杂交系统筛选与 H4相互作用的蛋白质.
목적:구건조단백 H2A、H2B、H3、H4적효모쌍잡교유이재체,검측기재효모세포중적표체화자격활활성,위응용효모쌍잡교계통사선여조단백상호작용적단백전정기출.방법:확증 H2A、H2B、H3、H4단백기편마구 cDNA서렬,극륭지효모표체재체 pGBK-T7중,재장기전화지효모세포 AH109중,제취효모총단백,검측각충조단백적표체,병검측표체적조단백재효모세포중대보고기유무자격활작용.결과:장 H2A、H2B、H3、H4기적 cDNA 극륭지효모표체재체 pGBK-T7중,기중조단백 H4득도정학표체,차대보고기 LacZ、HIS3、Ade 무격활작용.결론:가이이용효모쌍잡교계통사선여 H4상호작용적단백질.
Objective: To construct the yeast two-hybrid bait expression plasmids of histone H2A, H2B, H3 and H4, and to detect their protein expression and autonomous activation activity in yeast cells. Methods: The coding regions of H2A, H2B, H3 and H4 were amplified and cloned into the pGBK-T7 yeast two-hybrid vector, and were transformed into yeast AH109 cells. The total yeast protein was then extracted and the expression of the above histone proteins were detected and their autonomous activation activities were examined. Results: The cod?ing regions of H2A, H2B, H3 and H4 were cloned into the pGBK-T7 vector successfully and histone H4 got ex?pressed and showed no autonomous activation to the reporter gene LacZ, HIS3 and Ade. Conclusion: The yeast two-hybrid system can be applied to screen the interacting proteins of histone H4.