蕙兰MADS基因APETALA1/FRUITFULL-like的克隆和时空表达特性

Molecular cloning and spatiotemporal expression of an APETALA1/FRUITFULL-like MADS-box gene from the orchid (Cymbidium faberi)

摘要:

为研究兰花成花转变及花发育的调控机理，利用反转录RT-PCR和RACE的方法，从蕙兰萼片中克隆出一个APETALA1/FRUITFULL-like (AP1/FUL-like) 基因，命名为CfAP11，GenBank登录号为JQ031272.1。该基因编码的氨基酸序列与MADS-box蛋白家族中类AP1/FUL亚家族中球花石斛FRUITFULL-like具有较高的同源性 (84％)，系统进化分析表明该蛋白的氨基酸序列与AP1/FUL转录因子亚家族中的蛋白聚为一类。生物信息学分析推测表明，该基因编码的蛋白具有MADS保守域和相对保守的K区，二级结构中a-螺旋所占比例较高 (58.97％)，三级结构与月季、水稻和水仙非常相似。相对荧光定量PCR分析结果表明：CfAP11在根中表达痕量，生殖期比营养期叶片中表达量低、盛花期比花蕾期花葶中表达量高，由此推测，CfAP11可能与蕙兰的成花诱导、花发育有关；并发现CfAP11在盛花期花葶和子房中表达量远高于其他组织，表明其可能以某种机制参与果实的形成过程。

Abstract:

In order to identify genes involved in floral transition and development of the orchid species, a full-length APETALA1/FRUITFULL-like (AP1/FUL-like) MADS box cDNA was cloned from Cymbidium faberi (C. faberi) sepals and designated as C. faberi APETALA1-like (CfAP11, JQ031272.1). The deduced amino acid sequence of CfAP11 shared 84% homology with a member of the AP1/FUL-like group of MADS box genes (AY927238.1, Dendrobium thyrsiflorum FUL-like MADS box protein 3 mRNA). Phylogenetic analysis shows that CfAP11 belonged to the AP1/FUL transcription factor subfamily. Bioinformatics analysis shows that the deduced protein had a MADS domain and a relatively conservative K region. The secondary structure of CfAP11 mainly consisted of alpha helices (58.97%), and the three-dimensional structure of the protein was similar to that of homologues in Roza hybrida, Oryza sativa and Narcissus tazetta. Real-time quantitative PCR (qRT-PCR) results reveal low levels of its mRNA in roots, lower levels in leaves during reproductive period than vegetative period, and higher levels in pedicels at full-blossom stage than at bud stage. These results suggest that CfAP11 is involved in floral induction and floral development. Additionally, we observed higher levels of CfAP11 expression in pedicels and ovaries than in other tissues during full-blossom stage, which suggests that CfAP11 may also be involved in fruit formation in certain mechanism.