摘要
为明确CtbZIP47在酵母油脂合成中的作用,通过采用在线软件对红花CtbZIP47蛋白进行生物信息学分析;采用PCR方法扩增CtbZIP47基因cDNA序列;采用qRT-PCR方法检测该基因的表达情况;选择同源重组的方法构建pCAMBIA1302-CtbZIP47植物表达载体,利用激光共聚焦显微镜观察其在细胞中的定位情况;同时构建pPICZαA-CtbZIP47酵母表达载体,测定酵母中的油脂含量。结果表明:CtbZIP47蛋白由129个氨基酸编码组成;根据qRT-PCR结果显示,CtbZIP47基因在种子成熟后期表达量逐渐升高;通过生物信息学分析该蛋白为亲水性蛋白,具有核定位信号;亚细胞定位结果显示其定位在细胞核中;成功构建重组酵母表达载体pPICZαA-CtbZIP47,测定转基因酵母菌株的油脂含量,其含量显著高于毕赤酵母菌株和转空载体的毕赤酵母菌株。可见CtbZIP47基因在酵母油脂合成过程中起作用。
To clarify the role of CtbZIP47 in yeast oil synthesis,bioinformatics analysis of safflower CtbZIP47 protein was conducted using online software.The cDNA sequence of CtbZIP47 gene was amplified by PCR method,and the expression of the gene was detected by qRT-PCR method.The pCAMBIA1302-CtbZIP47 plant expression vector was constructed by homologous recombination method,and its localization in cells was observed by laser confocal microscopy.The yeast expression vector pPICZαA-CtbZIP47 was constructed to determine the lipid content in yeast.The experimental results showed that CtbZIP47 protein consists of 129 amino acids.The expression of CtbZIP47 gene was gradually increased in late stage of seed maturation by qRT-PCR detection.Bioinformatics analysis showed that CtbZIP47 protein is a hydrophilic protein with a nuclear localization signal,and it is localized in the nucleus.The recombinant yeast expression vector pPICZαA-CtbZIP47 was successfully constructed,and the oil content of transgenic strains was determined,which was significantly higher than that of the B.aeruginosa strains,indicating that CtbZIP47 gene plays a role in yeast lipid synthesis.
作者
刘怡麟
王帅
朱俐铭
王一凡
杨晶
LIU Yilin;WANG Shuai;ZHU Liming;WANG Yifan;YANG Jing(College of Life Sciences,Jilin Agricultural University,Changchun 130118,China;Engineering Research Center of Bioreactor and Pharmaceutical Development,Ministry of Education,Changchun 130118,China)
出处
《吉林农业大学学报》
CAS
CSCD
北大核心
2024年第3期431-438,共8页
Journal of Jilin Agricultural University
基金
国家自然科学基金青年基金项目(31801396)
吉林省教育厅科学技术研究项目(JJKH20200319KJ)。
关键词
红花
bZIP转录因子
毕赤酵母
油脂
safflower
bZIP transcription factor
Pichia pastoris
grease
