Bacillus sp.ZJS3基因组测序及砷胁迫下相关基因表达分析

Genome sequencing and expression of related genes of Bacillus sp. ZJS3 under As3+ stress

摘要:

【背景】环境中高毒性As³⁺的微生物氧化在砷的生物地球化学循环中起重要作用，具有潜在的应用价值。【目的】Bacillus sp.ZJS3菌株是本实验室前期分离鉴定的一株As³⁺耐受菌株，而且对多种重金属具有耐受性，期望进一步明确该菌株在As³⁺胁迫下菌体形态变化及应对砷胁迫的遗传基础，为As³⁺耐受细菌的研究提供基础数据。【方法】使用单分子实时测序（single-molecule real-time sequencing，SMRT）及Illumina测序技术对Bacillus sp.ZJS3菌株进行全基因组测序，对其基因进行功能注释和生物信息学分析，并结合绝对定量PCR技术对砷抗性及砷代谢相关基因进行分析。【结果】Bacillus sp.ZJS3菌株基因组大小为5.82 Mb，GC含量为35.9%，包含染色体1个、质粒3个、CDS数量为5 981个、tRNA 104个、sRNA 136个、rRNA 42个、串联重复序列173个、基因岛13个、转运蛋白1 023个、跨膜蛋白1 717个和双组分调控基因160个。NR、Swiss-Prot、Pfam、COG、GO和KEGG数据库分别可注释Bacillus sp.ZJS3菌株基因组中97.66%、69.30%、78.52%、65.49%、67.65%和43.87%的基因。绝对定量PCR结果表明，arsC基因在砷处理条件下显著高于对照组，而arsB基因在砷处理条件下显著低于对照组。【结论】Bacillus sp.ZJS3菌株在As³⁺胁迫下可能导致细胞分裂无法正常进行，进而影响细胞形态。基因组中aqpZ、arsA、arsB、arsC等基因的存在表明该菌株具有As³⁺外排和还原As⁵⁺的能力，phoUpstBACS的存在表明菌株可以吸收As⁵⁺，但菌株受到外界环境As³⁺胁迫时arsB表达水平降低。

Abstract:

[Background] Microbial oxidation of highly toxic As³⁺ in the environment plays an important role in the biogeochemical cycle of arsenic, which has potential application value.[Objective] Bacillus sp. ZJS3 has been identified to be tolerant to As³⁺, among other heavy metals. This paper aims to clarify the morphological changes of ZJS3 in response to As³⁺stress and the genetic basis for arsenic response, which is expected to provide basic data for research on As³⁺-tolerant bacteria. [Methods] The whole genome of ZJS3 was sequenced by single-molecule real-time (SMRT) sequencing and Illumina, followed by functional annotation and bioinformatics analysis of the genes. Genes related to arsenic resistance and arsenic metabolism were analyzed by absolute quantitative PCR. [Results] The genome of ZJS3 was 5.82 Mb, which contained 1 chromosome and 3 plasmids, with GC content of 35.9%, 5 981 coding sequences (CDSs), 104 tRNA genes, 136 sRNA genes, 42 rRNA genes, 173 tandem repeats, 13 gene islands, 1 023 transporter-coding genes, 1 717 transmembrane protein-coding genes, and 160 two-component regulatory genes. A total of 97.66%, 69.30%, 78.52%, 65.49%, 67.65%, and 43.87% of the genes in the genome of ZJS3 were annotated in NR, Swiss-Prot, Pfam, COG, GO and KEGG, respectively. arsC expression was significantly higher in the arsenic treatment group than in the control group, while the level of arsB was significantly lower in the arsenic treatment group than in the control group. [Conclusion] Under As³⁺ stress, they block cell division, further influencing cell morphology. The presence of aqpZ, arsA, arsB and arsC in ZJS3 genome indicates that the strain has the ability of As³⁺ efflux and As⁵⁺ reduction, and the presence of phoUpstBACS suggests that the strain can absorb As⁵⁺. However, the expression ofarsB decreases in response to external As³⁺ stress.