核桃JrGI基因克隆及表达分析

Cloning and expression analysis of JrGI gene in walnut

摘要:

GI （GIGANTEA）基因是生物节律钟关键输出基因，克隆核桃JrGI基因，并分析其在不同组织及不同时间的雌花芽表达情况，旨在为研究核桃JrGI基因的功能奠定基础。采用RT-PCR （reverse transcription-polymerase chain reaction）技术从‘新新2号’叶片中克隆获得JrGI基因全长序列，对其进行生物信息学分析、烟草亚细胞定位及表达分析。结果表明，JrGI基因全长为3 516 bp，编码1 171个氨基酸，分子量为128.60 kDa，等电点（isoelectric point）为6.13，属于亲水性蛋白；系统进化分析表明JrGI蛋白与胡杨GI蛋白亲缘关系最近。烟草亚细胞定位显示JrGI蛋白位于细胞核中。对‘新新2号’雌花芽不同时间段JrGI、JrCO和JrFT基因进行RT-qPCR （real-time quantitative PCR）分析，结果表明在形态分化临界期的雌花芽中表达量最高，推测JrGI在此时间对雌花芽的分化起到重要作用。另外，JrGI基因在‘新新2号’各组织中均有表达，且在叶片中表达量最高，推测JrGI基因在核桃叶片发育过程中也发挥重要功能。

Abstract:

GI (GIGANTEA) is one of the output key genes for circadian clock in the plant. The JrGI gene was cloned and its expression in different tissues was analyzed to facilitate the functional research of JrGI. RT-PCR (reverse transcription-polymerase chain reaction) was used to clone JrGI gene in present study. This gene was then analyzed by bioinformatics, subcellular localization and gene expression. The coding sequence (CDS) full length of JrGI gene was 3 516 bp, encoding 1 171 amino acids with a molecular mass of 128.60 kDa and a theoretical isoelectric point of 6.13. It was a hydrophilic protein. Phylogenetic analysis showed that JrGI of ‘Xinxin 2’ was highly homologous to GI of Populus euphratica. The result of subcellular localization showed that JrGI protein was located in nucleus. The JrGI, JrCO and JrFT genes in female flower buds undifferentiated and early differentiated of ‘Xinxin 2’ were analyzed by RT-qPCR (real-time quantitative PCR). The results showed that the expression of JrGI, JrCO and JrFT genes were the highest on morphological differentiation, implying the temporal and special regulation of JrGI in the differential process of female flower buds of ‘Xinxin 2’. In addition, RT-qPCR analysis showed that JrGI gene was expressed in all tissues examined, whereas the expression level in leaves was the highest. It is suggested that JrGI gene plays a key role in the development of walnut leaves.