南极适冷菌Psychrobacter sp.G假定蛋白基因PSYCG_10180对温度和盐度胁迫的应答特征分析  

Analysis of the Response of Hypothetical Protein Gene PSYCG_10180 to Temperature and Salinity Stress in the Antarctic Psychrotrophic Bacterium Psychrobacter sp.G

作　　者：张良[1,2] 李阳[1,2] 张政 林学政[1,2] 

机构地区：[1]国家海洋局第一海洋研究所,山东青岛266061 [2]国家海洋局海洋生物活性物质重点实验室,山东青岛266061

出　　处：《海洋科学进展》2017年第4期547-558,共12页Advances in Marine Science

基　　金：国家自然科学基金项目--南极适冷菌Psychrobacter sp.G温度与盐度胁迫下基因表达谱分析及冷/热激基因应答机制研究(41176174);南北极环境综合考察与评估专项--北极海域海洋生物和生态考察(CHINARE2016-03-05)和北极微生物资源多样性与功能评估(CHINARE2016-04-03)

摘　　要：根据南极适冷菌Psychrobactersp.G的基因组完成图,对其假定蛋白基因PSYCG_10180全长进行了基因克隆、序列分析与异源表达,并利用qRT-PCR和western blotting技术对其在不同温度/盐度胁迫条件下的表达特征进行了研究。序列分析表明,与蛋白PSYCG_10180相似性较高的均为假定蛋白;相似性最高的已知功能蛋白为蓝光传感蛋白(WP_010201745.1),二者的相似性仅有37%。在mRNA水平上,基因PSYCG_10180的表达被低温(0℃)显著诱导,在2,12h均被不同盐度显著抑制;在温度/盐度协同胁迫下,2h内低温均能诱导其表达,而高温低盐(30℃,15)则抑制其表达。在蛋白水平上,不同温度胁迫下,蛋白PSYCG_10180的表达量在2h变化不大,12h时均被诱导;在不同盐度胁迫下,该蛋白的表达在12h均被诱导;在温/盐协同胁迫下,除在2h被高温低盐(30℃,15)诱导外,其余条件下该蛋白的表达均被抑制。比较分析表明,在mRNA水平上,基因PSYCG_10180的表达更容易受温度的影响;在蛋白水平上,盐度对该基因表达的影响则要大于温度;在温盐协同胁迫时,盐度起主导作用。冻融试验表明,表达该假定蛋白可显著提高重组菌株的冻融存活率。本研究表明,假定蛋白基因PSYCG_10180可能在菌株Psychrobacter sp.G的低温等环境适应性中起着重要作用。According to the complete genome sequence,a hypothetical protein gene PSYCG_10180 was cloned from the Antarctic psychrotrophic bacterium Psychrobacter sp.G and it sequences was analyzed.The expression characteristics of gene PSYCG_10180 under different temperature and salinity stresses were investigated by quantitative real-time PCR（qRT-PCR）and western blotting.Sequence analysis showed that the proteins with relatively high similarity to PSYCG_10180 were unknown proteins,and the highest similarity of the known functional proteins（blue light sensor protein,WP_010201745.1）was only 37%.At the mRNA level,the expression of hypothetical gene PSYCG_10180 was induced by low temperature（0℃）;under different salinity stresses,the expression was inhibited at 2 hand 12 h;in the combined stress treatment,the expression was induced by low temperature at 2 h,and it was inhibited by high temperature,low salinity（30 ℃,15）.At the protein level,the expression level had little changes at 2 h,and it was induced at 12 hupon temperature stress investigated;in the combined stress treatment,the expression was inhibited except induced at 2 hby high temperature and low salinity（30℃,15）.Comparative analysis showed that,at the mRNA level,the expression of gene PSYCG_10180 was more sensitive to high temperature;at the protein level,the expression of hypothetical protein PSYCG_10180 was more sensitive to high salinity;in the combined stress treatment,the effect of salinity was dominative.Freezing and thawing test showed that the survival rate of the recombinant strain expressing the hypothetical protein was significantly enhanced compared with the control group.The study indicates that hypothetical protein PSYCG_10180 may play an important role in environmental adaptability of Psychrobacter sp.G.

关 键 词：PSYCHROBACTER PSYCG10180 胁迫表达 QRT-PCR WESTERN BLOTTING 

分 类 号：Q751[生物学—分子生物学]