DELLA蛋白GbGAI4在海岛棉体细胞胚发生过程中的功能机制研究

海岛棉是新疆重要的农业经济作物。目前高效体细胞胚体系的建立困难严重地制约着海岛棉分子育种进程。本研究围绕DELLA蛋白基因GbGAI4介导的海岛棉体胚发生体系中由体细胞退分化为胚性细胞的瓶颈环节的功能机制为核心，拟从以下几方面研究：（1）利用启动子GUS活性检测和实时定量PCR技术检测GbGAI4在海岛棉胚性细胞分化过程中的表达模式和启动子特性；（2）借助激光共聚焦显微镜技术和Western Blot技术检测GbGAI4蛋白的亚细胞定位和蛋白稳定性；（3）通过酵母双杂交等技术筛选并验证胚性细胞分化过程中GbGAI4蛋白的互作蛋白；（4）借助生长发育特异性启动子控制GbGAI4和生长发育调控基因组合（BBM/LEC/WUS）在细胞中的表达，初步鉴定GbGAI4在海岛棉胚性细胞分化过程中的生理作用。本研究的实施有助于拓展海岛棉体细胞胚发生体系的基因型，从而加快新疆海岛棉分子育种进程。

Gossypium barbadense, as Xinjiang important economic crop, faces with the challenge of cotton variety improvement. However, the difficulties of establishment of an efficient somatic embryogenesis system restrict the molecular breeding process in G. barbadense. For the bottleneck in the dedifferentiation process from somatic cell into embryonic cell in G.barbadense embryogenesis, this project intends to study the functions and molecular mechanism of GbGAI4 from the following aspects: (1) analysis the expression pattern and promoter characteristics of GbGAI4 in G. barbadense somatic embryogenesis with real-time PCR and promoter GUS-activities assay; (2) check the subcelluar localization and stability of GbGAI4 through the confocal laser microscopy technology and western blot; (3) screen and verify the GbGAI4-interacted proteins with the yeast two-hybrid assay and the other technologies; (4) Preliminarily elucidate the physiological roles of GbGAI4 in the embryonic-cell differentiation process through controlling the expressions of GbGAI4 and the development regulator genes combination (BBM/LEC/WUS) with the development-specific promoter in cotton cells. The implementation of the project contributes to expand the genotypes of G.barbadense with somatic embryogenesis, which promote the molecular breeding process of G.barbadense in Xinjiang.