Transcriptome profile of Haloferax volcanii H26 strain under sodium bisulfite induction (RNA-seq)

DNA deamination occurs constantly in a cell and causes DNA damage. As this damage can be deleterious, organisms have evolved many systems to eliminate it. Deamination of cytosine, guanine, adenine, and 5-methylcytosine results in the formation of uracil, xanthine, hypoxanthine, and thymine, respectively. Sodium bisulfite is a kind of DNA deaminating agent that can increase the frequency of DNA deamination in cells. This study measures the transcriptome profile of Haloferax volcanii H26 strain and HVO_RS06830 gene knockout strain, induced with different concentrations of sodium bisulfite. Overall design: Haloferax volcanii H26 strain and HVO_RS06830 gene knockout strain were grown in Hv-YPC medium at 45°C until OD600 reach 0.5. Subsequently, sodium bisulfite was added to the final concentration of 0.1 or 0.5 mM and continuously cultured for 48 h. Cells were harvested by centrifugation at 12,000 rpm at 4°C for 20 min. After that, total RNA was extracted immediately using Direct-zol RNA MiniPrep Kits. Sequencing libraries were generated using NEBNext Ultra II Directional RNA Library Prep Kit for Illumina following manufacturer's recommendations, and sequenced by Illumina NovaSeq6000 PE150 platform. The experiment was repeated two times.

DNA脱氨（DNA deamination）在细胞内持续发生并引发DNA损伤。鉴于此类损伤具有有害性，生物体已演化出多种系统以清除该损伤。
胞嘧啶、鸟嘌呤、腺嘌呤及5-甲基胞嘧啶的脱氨反应，可分别生成尿嘧啶、黄嘌呤、次黄嘌呤与胸腺嘧啶。
焦亚硫酸钠（Sodium bisulfite）是一类DNA脱氨试剂，能够提升细胞内DNA脱氨的发生频率。本研究对经不同浓度焦亚硫酸钠诱导的火山嗜盐盒菌（Haloferax volcanii）H26菌株与HVO_RS06830基因敲除菌株的转录组谱进行了检测。
实验整体设计：将火山嗜盐盒菌H26菌株及HVO_RS06830基因敲除菌株置于Hv-YPC培养基中，于45℃条件下培养至OD600达到0.5。随后向培养基中加入焦亚硫酸钠，使其终浓度分别为0.1 mM或0.5 mM，继续培养48小时。以4℃、12000 rpm离心20分钟收集细胞，随即使用Direct-zol RNA MiniPrep试剂盒提取总RNA。按照厂商推荐的操作流程，使用NEBNext Ultra II Directional RNA Library Prep Kit for Illumina构建测序文库，并通过Illumina NovaSeq6000 PE150平台完成测序。本实验共重复两次。