高羊茅FaCCA1基因克隆、表达分析及基因编码蛋白的亚细胞定位  

Cloning and expression analysis of FaCCA1 gene and its protein subcellular localization in tall fescue

作　　者：陈锡[1,2] 陈莹[1] 刘晓霞 舒健虹 王小利[1] 赵德刚[1,2] CHEN Xi;CHEN Ying;LIU Xiaoxia;SHU Jianhong;WANG Xiaoli;ZHAO Degang(Guizhou Institute of Prataculture,Guizhou Academy of Agricultural Sciences,Guiyang 550006,China;College of Life Sciences,the Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region,Ministry of Education,the National-local Joint Engineering Research Center of Karst Region Plant Resources Utilization&Breeding,Guizhou University,Guiyang 550025,China)

机构地区：[1]贵州省农业科学院/贵州省草业研究所,贵阳550006 [2]贵州大学生命科学学院/山地植物资源保护与种质创新教育部重点实验室/喀斯特山区植物资源利用与育种国家地方联合工程研究中心,贵阳550025

出　　处：《植物生理学报》2021年第9期1767-1777,共11页Plant Physiology Journal

基　　金：国家自然科学基金(31860674);贵州省科技计划([2019]1302)。

摘　　要：生物钟节律基因(circadian clock gene)CCA1在植物光周期途径中具有十分重要的作用。以高羊茅‘黔草1号’cDNA为模板,利用同源扩增结合cDNA末端快速扩增(rapid amplification of cDNA ends,RACE)技术从高羊茅中获得与拟南芥CCA1高度同源的cDNA全长序列,命名为FaCCA1。利用BLAST在线软件对该基因进行生物信息学分析,通过农杆菌介导的烟草瞬时表达,观察Fa CCA1基因编码蛋白的亚细胞定位,并利用RT-qPCR技术分析不同光照条件下FaCCA1基因表达水平。结果表明,该序列cDNA全长为2433bp,含有一个完整的开放阅读框,长度为2148 bp,编码蛋白含有715个氨基酸,蛋白分子量为77.80 kDa,理论等电点为5.91。亚细胞定位结果显示FaCCA1基因所编码的蛋白定位于细胞核。系统进化树分析表明FaCCA1与禾本科植物的CCA1蛋白亲缘关系较近,其中与大麦亲缘关系最近,达到80%以上。FaCCA1基因表达量在长日照高于短日照,表达水平出现昼夜节律钟变化模式,表明高羊茅FaCCA1表达量受光周期调控。这为高羊茅FaCCA1基因的功能研究提供参考资料,为高羊茅的分子育种利用提供了改良靶标。Circadian clock gene CCA1 plays a very important role in photoperiodic pathway of plant. The sequence of Fa CCA1 gene is highly homologous with CCA1 of Arabidopsis thaliana and was cloned by homologous amplification and rapid amplification of cDNA ends(RACE) from cDNA of Festuca arundinacea ‘Qiancao No.1’. Its bioinformatics was analyzed by BLAST, the subcellular localization of FACCA1 gene encoding protein was observed by Agrobacterium tumefaciens-mediated transient expression in tobacco, and the expression level of FaCCA1 was analyzed by RT-qPCR under different light conditions in tall fescue.The full-length cDNA(2 433 bp) of FaCCA1 was obtained with a predicated 2 148 bp open reading frame, which encoded a protein containing 715 amino acids. Relative molecular weight of FaCCA1 protein was 77.80 kDa, and its theoretical isoelectric point was 5.91. FaCCA1 was located on the cell nucleus. Phylogenetic analysis showed that FaCCA1 was closely related with the CCA1 protein of gramineous plant, in which FaCCA1 was most closely related to barley(more than 80% homology). Expression level of FaCCA1 was higher in long-day than that in short-day treatments, indicating that expression level of FaCCA1 is regulated by photoperiod. This study can provide basic data to study function of the FaCCA1 gene in the future and a target gene for molecular breeding in tall fescue.

关 键 词：高羊茅 FaCCA1 亚细胞定位 表达分析 

分 类 号：S688.4[农业科学—观赏园艺]