金边红苞凤梨叶片的超微观察和AbGLK1的克隆与表达分析

Ultrastructure of Leaf and Cloning, Expression of AbGLK1 in Ananas comosus var. bracteatus 'Chiyan'

摘要:

为了解Golden2-like (GLK)转录因子在金边红苞凤梨(Ananas comosus var. bracteatus ‘Chiyan’)绿白嵌合叶片形成中的作用，采用RT-PCR技术克隆得到了AbGLK1基因, 其开放阅读框全长1 371 bp，编码456个氨基酸，含有1个GARP-DNA结合域和1个C末端结构域GCT box (GOLDEN2 C-terminal box)，属于GLK转录因子家族，在酵母中具有转录因子的转录激活活性。烟草亚细胞定位表明AbGLK1蛋白定位在细胞核。RT-qPCR分析表明，AbGLK1基因在金边红苞凤梨的根、茎、叶中均有表达，但具有组织器官差异性，在叶片中的表达量显著高于根和茎(P < 0.05)。AbGLK1基因在叶片边缘白化组织中的表达量显著低于绿色组织，约为绿色组织的1/3 (P < 0.05)。叶片白化组织叶绿体内膜系统模糊，无类囊体存在，含有大量囊状小泡，质体小球数量多且体积较大。因此，推测AbGLK1基因可能参与了金边红苞凤梨中的叶绿体发育，其下调表达可能导致叶片白化组织中叶绿体发育不成熟。

关键词:金边红苞凤梨;叶绿体;超微结构;AbGLK1;基因克隆;转录激活活性;基因表达

Abstract:

In order to understand the function of Golden2-like (GLK) in green-white chimeric leaf formation of Ananas comosus var. bracteatus 'Chiyan', AbGLK1 gene was cloned by RT-PCR. The results showed that AbGLK1 contained an open reading frame with 1 371 bp, encoding a protein with 456 amino acids. AbGLK1 protein had a GOLDEN2 C-terminal box and a GAPR DNA-binding domain at the N terminal, belonged to GLK transcription factor family, which located in the nucleus. Transactivation analysis showed that AbGLK1 had transcriptional activation activity. And AbGLK1 was tissue-specific expressed in root, stem and leaf of 'Chiyan'. The expression of AbGLK1 in leaves was significantly higher than that in roots and stems (P < 0.05); which in albino tissues was about 1/3 in green tissues (P < 0.05). In albino tissue, there was no thylakoid in chloroplast, but large number of plastoglobulus and vesicles exist in chloroplast. Therefore, it is suggested that AbGLK1 might be involved in chloroplast development, and down expression of AbGLK1 might lead to immature chloroplast in albino tissue.

Key words:Ananas comosus var. bracteatus 'Chiyan';Chloroplast;Ultrastructure;AbGLK1;Gene cloning;Transcriptional activation;Gene expression