文心兰COL基因的分离及其表达分析

Isolation and Expression Analysis of COL Genes of Oncidium

CONSTANS(CO)及CONSTANS-like(COL)基因在光周期调控植物开花中起到重要的作用。该研究以文心兰(Oncidium)品种‘金辉’为材料,分离了CO同源基因OnCOL2及另外2个COL基因(OnCOL8和OnCOL9),它们分别编码326、411和291个氨基酸;生物信息分析预测它们均定位于细胞核;OnCOL2、OnCOL8有2个锌指B-box结构域和1个CCT结构域,而OnCOL9缺少B-box结构域。多序列比对及进化树分析结果表明,所有COL蛋白可划分为3组,OnCOL2与OnCOL8、OnCOL9分到了2个不同组中。OnCOL2与建兰(Cymbidium ensifolium)CeCOL(90.77%)高度相似;OnCOL8与OnCOL9在进化关系上更为接近,分别与拟南芥(Arabidopsis thaliana)AtCOL9和AtCOL10关系最近,它们在B-box和CCT结构域都极为保守。表达分析结果表明OnCOL2、OnCOL8与OnCOL9分别在花、根和假鳞茎中表达量最高,在叶片中的表达呈周期性变化趋势,且在长、短日照条件下表达的峰值及时间均存在差异,在花芽分化时期的叶片中表达量均显著上调。研究表明,OnCOL2、OnCOL8与OnCOL9基因均受到生物钟和日长调节,在光周期途径中,可能通过上调它们的表达以促进文心兰花芽的形成。该研究结果为进一步研究基因功能及光周期调控文心兰开花机制奠定了基础。

The CONSTANS(CO)and CONSTANS-like(COL)genes play a key role in photoperiod regulation of flowering.In this study,A CO ortholog,OnCOL2,and two COL paralogs(OnCOL8 and OnCOL9)were isolated from Jinhui,a variety of Oncidium,encoding 326,411 and 291 amino acids,respectively.The bioinformatics analysis showed that they were predictively unstable proteins and located in the nucleus.OnCOL2 and OnCOL8 contained two B-box domains of Zinc finger and one CCT domain,while OnCOL9 lacked B-box domain.The multiple sequence alignment and phylogenetics analysis showed that all COLs could be divided into three groups,and OnCOL2,OnCOL8 and OnCOL9 were divided into two different groups.OnCOL2 shared highly identity with Cymbidium ensifolium CeCOL(90.77%).OnCOL8 and OnCOL9 were closer in evolutionary relationship,and exhibited the highest similarity with AtCOL9 and AtCOL10 in Arabidopsis,respectively.They were extremely conservative in the B-box and CCT domains.Expression analysis showed that OnCOL2,OnCOL8 and OnCOL9 had the highest expression in flowers,roots and pseudobulbs,respectively.Their expressions in leaves showed a periodic trend,and the peak values and time were different under long-day and short-day conditions.In the flower bud differentiation stage of Oncidium,their expressions were significantly up-regulated in leaves.These results indicated that OnCOL2,OnCOL8 and OnCOL9 were regulated by circadian clock and day length,and their expression up-regulated could probably promote the flower bud formation of Oncidium in the photoperiod pathway.The results provide the basis for further study on gene function and photoperiod regulation of flowering mechanism of Oncidium.