芽殖酵母Sik1蛋白在纺锤体定位中的功能研究
Functional study of the Saccharomyces cerevisiae Sik1 protein in spindle orientation
投稿时间:2020-12-25  修订日期:2021-02-23
DOI:
中文关键词: 芽殖酵母  纺锤体定位  Sik1  Num1  核迁移
英文关键词: Budding yeast  Spindle orientation  Sik1  Num1  Nuclear migration
基金项目:国家自然科学基金青年基金项目;中南民族大学基本科研业务费专项资金项目
作者单位E-mail
唐仙英 中南民族大学 xytangscuec@163.com 
庞文颖 中南民族大学生命科学学院 1973615713@qq.com 
海力 中南民族大学生命科学学院  
滕雪 中南民族大学生命科学学院  
张楠 中南民族大学生命科学学院  
朱顺 中南民族大学生命科学学院  
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中文摘要:
      为了探究芽殖酵母蛋白Sik1在有丝分裂纺锤体定位中的功能,利用同源重组技术将SIK1的内源启动子替换成GAL1启动子,得到基因组位点过表达SIK1的菌株,同时使菌株表达GFP-Tub1,便于观察纺锤体;接着在12 ℃低温条件下培养细胞以促进双核细胞的产生,细胞经乙醇固定、DAPI染色后在荧光显微镜下观察细胞核分裂及纺锤体定位;最后检测SIK1过表达对细胞生存能力的影响. 结果表明:过表达Sik1干扰纺锤体定位,但对细胞核迁移没有明显影响. 在16℃低温胁迫条件下,过表达Sik1降低了纺锤体定位突变体kar9Δ的生长活力却部分恢复了num1Δ细胞的生长,说明Sik1可能在Dynein通路中起作用.
英文摘要:
      To explore the function of Saccharomyces cerevisiae Sik1 protein in spindle orientation, the endogenous promoter of SIK1 was replaced with the GAL1 promoter through homologous recombination approach to obtain the strain that overexpresses SIK1 in the chromosomal locus. The strain was also constructed to express GFP-Tub1 to observe spindles. Next, cells were incubated at 12℃ to facilitate the production of bi-nucleated cells. Cells were fixed with ethanol, stained with DAPI, and observed with fluorescence microscope to examine nuclear division and spindle orientation. Finally, effect of overexpressing Sik1 on cell viability was examined. The results showed that overexpressing Sik1 disturbed spindle orientation, but had no evident negative effect on nuclear migration. Under the stress of low temperature 16 ℃, overexpression of Sik1 reduced the viability of kar9Δ, a spindle orientation mutant, but recovered the viability of num1Δ cells partially, suggesting that Sik1 might play a role in the Dynein pathway.
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