油茶CoPIF3基因的克隆及表达分析

Cloning and expression analysis of CoPIF3 gene in Camellia oleifera

本研究以油茶品种‘华硕’为材料,通过RT-PCR克隆得到光敏色素互作因子中的一条PIF3基因,命名为CoPIF3。CoPIF3基因序列全长1884 bp,编码627个氨基酸。CoPIF3蛋白分子量为67.881 kDa,属于bHLH蛋白家族,该基因与茶树的PIF3亲缘关系较近,被定位在细胞核中,并成功在E.coli Rosetta(DE3)菌株中表达。实时荧光定量表明:CoPIF3基因在叶芽及老茎中的表达量显著高于其他部位;在自交授粉下,CoPIF3基因在子房中24 h的表达量最低,在花柱中表达量在授粉后上调至24 h时最高;在异交授粉下,CoPIF3基因在子房和花柱中的表达量分别在24 h和48 h时达到峰值,随后均下调。研究表明CoPIF3可能参与了油茶自交授粉后花粉管的生长,进而参与了油茶自交不亲和反应。本研究为油茶自交不亲和机制的解析提供参考。

In this study,a member of phytochrome-interacting factors named CoPIF3 was cloned by RT-PCR from Camellia oleifera’Huashuo’.The ORF of the gene was 1884 bp and encoded 627 amino acids.CoPIF3 protein had a molecular weight of 67.881 kDa and belonged to the bHLH protein family,this gene was closely related to PIF3 of tea tree.It was localized in the nucleus and successfully expressed in E.coli Rosetta(DE3)strain.Quantitative real-time PCR results showed that the expression of CoPIF3 gene in leaf buds and old stems was significantly higher than in other parts.In self-pollination,the expression of CoPIF3 was the lowest in the ovary at 24 h,and its expression in the style was up-regulated at 24 h after pollination.In cross-pollination,the expression of CoPIF3 gene in ovary and style peaked at 24 h and 48 h,respectively,and then down-regulated.It is suggested that CoPIF3 may be involved in the growth of pollen tubes after pollination,and then participate in self-incompatibility of C.oleifera.This study provide reference for the analysis of the self-incompatibility mechanism of C.oleifera.