【摘要】 目的研究耐辐射奇球菌Deinococcus radiodurans R1基因组DNA酶切的优化条件,从而获得构建基因文库所需的DNA片段,旨在构建DR菌基因组DNA表达文库,进一步筛选文库中与其有相互作用的蛋白。方法培养耐辐射奇球菌,提取基因组DNA,用Sau3AI酶切DR菌基因组DNA,分别从酶浓度、酶切时间等选择酶切产生的DNA片段集中在0.5~5.0 kb的最佳条件,最后通过琼脂糖凝胶电泳进行检测。结果Sau3AI酶切DR菌基因组DNA的最佳酶浓度为0.125 u/10μL,最佳作用时间为3 h,此条件下DR菌基因组DNA片段主要集中于0.5~5.0 kb。结论优化了DR菌基因组DNA的酶切条件,为进一步构建DR菌基因组DNA表达文库,筛选与高抗辐射相关基因产物的互作蛋白奠定了基础。更多还原

【Abstract】 Objective To construct the genomic DNA expression library and then screen to isolate proteins that might interact with the anti-radiation related proteins by the yeast two-hybrid system,the condition of digestion for Deinococcus radiodurans R1 genome was optimized. Methods The Deinococcus radiodurans R1 was cultivated and the genomic DNA was extracted,the digestion conditions including enzyme concentration and reaction time with Sau3AI for Deinococcus radiodurans R1 genome were optimized.Finally the results were checked by agarose gel electrophoresis. Results The results showed that the best Sau3A1concentration and reaction time for DR genome DNA are 0.125 U/10 μL and 3 hours.Conclusions The digestion condition for DR genome has been optimized,which lays a foundation for constructing the genomic DNA expression library and further screening for the proteins that might interact with the anti-radiation related proteins from Deinococcus radiodurans R1.更多还原