棉花GhMADS29启动子克隆及表达分析

Cloning and Expression Analysis of the Promoter of GhMADS29 from Cotton

以实验室克隆的GhMADS29(GeneBank登录号:JQ682642)基因的cDNA序列Blast搜索雷蒙德氏棉的基因组序列,根据Blast结果设计引物,克隆到起始密码子上游-19位开始的1316 bp的序列;利用PlantCARE启动子在线分析软件预测其含有核心启动子元件TATA-box和CAAT-box,并含有光、温、赤霉素、水杨酸、生长素等的响应元件。通过替换pBI121载体上的CaMV35S启动子构建了GhMADS29启动子与GUS基因的融合表达载体并转化拟南芥,组织化学染色分析发现其在14 d幼苗的根和叶中都有表达,在萼片、花瓣、雌蕊、果瓣中也表达,而在雄蕊和种子中不表达。综上所述,我们推测GhMADS29可能与各种开花途径有关,与萼片、花瓣、雌蕊等花器官的发育有关,还可能与果实是否开裂有关。

We used the cDNA sequence of GhMADS29 as a search query to BLAST the genome sequences of Gossypium rai- mondii L. We designed the primers to clone the promoter according to the BLAST results. We obtained a 1316-bp sequence up- stream from the -19 site of the start codon. Sequence analysis showed that this promoter region contained a TATA-box core ele- ment, a CAAT-box, and other cis-acting elements, including an AuxRR core, a TCA element, TC-rich repeats, an HSE, Box I, a GARE motif, and a TGA element; these elements are involved in auxin, salicylic acid, defense and stress, heat, light, and gib- berellin responses, respectively. We replaced the CaMV 35S promoter in the pBI121 vector with the 1316-bp promoter sequence to construct a fusion vector containing the GhMADS29 promoter connected to the GUS gene. The fusion vector was designated as pSVP121. GUS histochemical staining analysis of transgenic Arabidopsis carrying the pSVP121 vector showed that the pro- moter was active in the roots and leaves of 14-day-old seedlings and in floral organs（sepals, petals, and pistils）. It also showed activity in silique valves, but not in stamens and seeds. These results suggest that GhMADS29 may be regulated by light, temper- ature, and hormones, and it may also participate in the development of sepals, petals, and pistils. Whether or not the siliques will be dehiscent may be related to the function of GhMADS29 because of the activity of its promoter in the silique valves.