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| 番茄SlCDJ2蛋白在大肠杆菌和酵母中的表达研究 |
| The study of tomato SlCDJ2 protein expression in Escherichia coli and yeast |
| 投稿时间：2018-11-10 修订日期：2019-02-18 |
| DOI： |
| 中文关键词: SlCDJ2蛋白;大肠杆菌;酵母;渗透胁迫 |
| 英文关键词: SlCDJ2 protein; E. coli; S. cerevisiae; osmotic stress |
| 基金项目:山东省自然科学基金博士基金(ZR2018BC032)；济宁医学院大学生创新项目(100220004030)；国家级大学生创新项目(201710443030)；济宁医学院博士基金(600492001) |
| 作者 | 单位 | 邮编 | | 刘琪 | 济宁医学院生物科学学院 山东日照 276826 | 276826 | | 商鑫涛 | 济宁医学院生物科学学院 山东日照 276826 | | | 陈晨 | 济宁医学院生物科学学院 山东日照 276826 | | | 王国栋 | 济宁医学院生物科学学院 山东日照 276826 | 276826 |
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| 中文摘要: |
| 目的 DnaJ蛋白作为一种广泛存在于细胞内的分子伴侣,能响应多种胁迫反应。本研究以转基因大肠杆菌和酿酒酵母为材料,研究番茄Solanum lycopersicum Chloroplast DnaJ 2(SlCDJ2)蛋白在渗透胁迫下的表达情况,为进一步在番茄中研究其功能提供初步的理论依据。方法 采用PCR的方法扩增获得SlCDJ2基因,构建大肠杆菌原核表达重组质粒pET30a-SlCDJ2,构建酵母真核表达重组质粒pYES2-SlCDJ2,分别通过热激转化法和醋酸锂酵母转化获得转基因大肠杆菌和酵母菌株。最后通过SDS-聚丙烯凝胶电泳分析SlCDJ2蛋白大肠杆菌中的诱导表达情况,采用分光光度法测定转基因大肠杆菌在渗透胁迫下的生长曲线,另外,通过涂布的方法来观察转基因酵母菌在渗透胁迫下的生长表型。结果 SDS-聚丙烯凝胶电泳结果显示,SlCDJ2蛋白在大肠杆菌中成功诱导表达,分光光度法测定结果表明,转基因大肠杆菌在渗透胁迫下吸光度明显高于野生型大肠杆菌。酵母菌的生长表型分析发现,转基因酵母菌在渗透胁迫下的生长菌落明显好于野生型酵母菌。结论 该研究表明,在大肠杆菌和酿酒酵母中过表达SlCDJ2基因,有助于提高大肠杆菌和酿酒酵母对渗透胁迫的抗性。 |
| 英文摘要: |
| Objective DnaJ protein, as a kind of molecular chaperone widely existing in cells, can respond to a variety of stress responses. In this study, transgenic Escherichia coli and Saccharomyces cerevisiae were used as materials to study the expression of tomato SlCDJ2 protein under osmotic stress, providing a preliminary theoretical basis for further studying its function in tomatoes. Methods The SlCDJ2 was obtained by PCR amplification, the recombinant plasmid pET30a-SlCDJ2 and pYES2-SlCDJ2 were constructed and transffered into E. coli and S. cerevisiae by heat shock and lithium acetate yeast transformation methods to abtain the transgenic E. coli and S. cerevisiae. Finally, SDS-polypropylene gel electrophoresis (SDS-PAGE) was used to analyze the expression of SlCDJ2 protein in E. coli, and the growth curve of transgenic E. coli under osmotic stress was measured by spectrophotometry.SIn addition, the growth phenotype of transgenic yeast under osmotic stress was observed by coating. Results SDS-PAGE results showed that SlCDJ2 protein was successfully induced in E. coli, and the results of spectrophotometric determination showed that the absorbance of transgenic E. coli was significantly higher than that of wild escherichia coli under osmotic stress. The growth phenotype analysis of yeast showed that the growth colony of transgenic yeast was better than that of wild yeast under osmotic stress. Conclusions The study showed that SlCDJ2 overexpression was helpful to improve the osmotic stress resistance of E. coli and S. cerevisiae. |
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