耐辐射奇球菌FMN riboswitch的抗氧化性初步研究.docVIP

  耐辐射奇球菌 FMN riboswitch 的抗氧化性 初步研究# 杨鹏，陈周伟，单战，刘立丽，丁先锋，郭江峰** 5 10 15 20 25 30 35 （浙江理工大学生命科学院，杭州 310018） 摘要：FMN riboswitch 是一种能调控黄素单核苷酸生物合成和转运基因的特异性 mRNA 非 编码结构，它能和小分子代谢物（FMN）或通过感知外界环境变化，同时不需要蛋白因子 的参与来调控基因的表达。利用体外重叠延伸 PCR 扩增技术和体内同源重组方法，获得了 耐辐射奇球菌（Deinococcus radiodurans）FMN riboswitch 全基因缺失突变株。对野生型和突 变株分别进行生长曲线的测定结果表明，突变株生长延缓，生长速度比野生型 R1 慢。对野 生型和突变株分别用不同浓度过氧化氢（H2O2）处理，结果表明，在 50 mM H2O2 的条件 下，与野生型相比（存活率为 56%），突变株存活率为 40%，说明 FMN riboswitch 的缺失 使耐辐射奇球菌对过氧化氢更敏感。荧光定量 QPCR 技术结果表明，在 4 h 内，耐辐射奇球 菌中 FMN riboswitch 表现为先上调后下调的趋势。以上结果表明，FMN riboswitch 参与耐辐 射奇球菌氧化抗性的形成，可能在高效的氧化保护体系中发挥重要的作用。 关键词：耐辐射奇球菌；FMN riboswitch；氧化抗性 中图分类号：Q936 Role of FMN Riboswitch in oxidative stability in Deinococcus radiodurans YANG Peng, CHEN Zhouwei, SHAN Zhan, LIU Lili, DING Xianfeng, GUO Jiangfeng (College of Life Science, Zhejiang Sci-Tech University, HangZhou 310018) Abstract: FMN riboswitch is structured noncoding RNA domains that control gene expression by selectively binding metabolites (FMN) or sensing surroundings changes without the need for protein factors, which is involved in the biosynthesis and transport of riboflavin (vitamin B2) and related compounds. In this work, we constructed the deletion mutant of FMN riboswitch to investigate the possible role of FMN riboswitch in response to oxidative stress in Deinococcus radiodurans. The growth curve showed that the deletion of FMN riboswitch resulted in an obvious growth delay and its growth is slower than that of D. radiodurans. Compared with the survival rate of 56% of D. radiodurans, the mutation strain survived under 50 mM of H2O2 with a survival rate of 40%, indicating that deletion of FMN riboswitch exhibited an obviously increased susceptibility to hydrogen peroxide. Furthermore, quantitative real-time PCR analysis demonstrated that, during the course of hydrogen peroxide treatment of 4 h, FMN riboswitch have a trend of over-expressed first and low-expressed thereafter. These results suggest that FMN riboswitch plays an important