文心兰OnRR10基因的克隆及表达分析

Cloning and Expression Analysis of OnRR10 Gene in Oncidium

为了鉴定控制文心兰花瓣衰老的关键基因,从文心兰花瓣中提取总RNA,利用RACE技术,得到OnRR10基因(登录号:MN337795)。该基因全长630 bp,编码209个氨基酸大小的蛋白质。蛋白质二级结构分析表明,OnRR10蛋白为亲水性蛋白,定位于细胞核中。同源序列比对及系统进化分析显示,OnRR10基因属于YesN家族。兰科中,OnRR10蛋白与小兰屿蝴蝶兰OnRR10亲缘关系最近。实时荧光定量分析表明,OnRR10基因在文心兰花发育的A时期(花苞期)和F时期(衰老早期)表达量达到最高值;基因表达似不受生物钟控制;细胞分裂素处理后表达分析表明,短时间内受6-BA刺激基因表达量急速上升。结果表明,OnRR10基因可能通过细胞分裂素作用而参与文心兰花瓣衰老过程。

In order to characterize the key genes which control the petal senescence,a gene encoding an response regulator(RR),OnRR10 gene(GenBank accession number:MN337795),was isolated from the petals of Oncidium with RACE(rapid amplification of cDNA ends).The gene is 630 bp in length and encodes a protein of 209 amino acids in size.Protein secondary structure analysis showed that the OnRR10 protein is a hydrophilic protein and is localized in the nucleus.Homologous sequence alignment and phylogenetic analysis showed that the OnRR10 belongs to the YesN family.In Orchidaceae,OnRR10 protein is closely related to the ORR10 gene of Phalaenopsis equestris.Real-time quantitative analysis showed that the expression level of the OnRR10 gene was the highest at the bud swelling stage and flower initial senescence stage of Oncidium during the flower development,and that the expression of the OnRR10 gene was not regulated by the circadian clock,but increased rapidly in a short time under the 6-BA(6-Benzylaminopurine)treatment.These results implicated that the OnRR10 gene may be involved in the petal senescence through cytokine effect.