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目的:构建耐辐射奇球菌(Deinococcus radioduransR1)基因组DNA表达文库,为进一步研究耐辐射奇球菌高抗辐射的调控网络奠定基础。方法:提取耐辐射奇球菌基因组DNA,用Sau3AI酶将基因组DNA部分酶切成0.5-5kb大小的片段,用T4DNA连接酶将部分酶切片段与经BamHⅠ和碱性磷酸酶(CIAP)处理的pGADT7载体进行连接后电击转化大肠杆菌DH5α。结果:得到重组子数为2.2×104,扩增后的文库滴度为108cfu/mL。结论:构建了耐辐射奇球菌基因组pGADT7表达文库,为进一步筛选与高抗辐射相关基因产物的互作蛋白奠定了基础。
OBJECTIVE: To construct a genomic DNA expression library of Deinococcus radiodurans R1, which lays the foundation for further study on the regulatory network of Radiation resistant Deinococcus radiodurans. Methods: The genomic DNA of D. radiodurans was extracted. The genomic DNA was partially digested with Sau3AI enzyme into 0.5-5kb fragment. The partially digested fragment was digested with T4 DNA ligase and ligated with pGADT7 treated with BamHI and alkaline phosphatase (CIAP) The vector was ligated and electroporated into E. coli DH5α. Results: The number of recombinants was 2.2 × 104, and the amplified library titer was 108 cfu / mL. Conclusion: The construction of the genome of M. radiodurans genome pGADT7 expresses the foundation for further screening of the interacting proteins with the gene products related to high radiation resistance.