《植物生理学报》 2015, 51(4): 517-522

剑麻AsKNOX I基因克隆及蛋白表达分析

陆军迎, 张燕梅, 李俊峰, 周文钊*

中国热带农业科学院南亚热带作物研究所, 海南省热带作物营养重点实验室, 广东湛江524091

通信作者：周文钊；E-mail: wenzhaozhou@163.com；Tel: 0759-2859216

摘　要：

植物的同源异型盒基因(knotted1-like homeobox, KNOX)与叶片形态建成具有密切的相关性。本研究以剑麻(Agave sisalana)茎顶端分生组织为材料分离得到编码357个氨基酸的AsKNOX I全长基因, 预测该蛋白的分子量约为40 kDa, 等电点为pI 6.4, 且包含KNOX I同源蛋白家族典型的HD、MEINOX、ELK和GSE保守结构域。后通过构建原核和真核表达载体, 分别在大肠杆菌和烟草叶片中瞬时表达, 用His标签进行蛋白纯化, Western blot用于表达产物检测。结果表明, AsKNOX I基因已在原核和真核中成功表达, 为进一步分析该蛋白的结构及功能特征提供了技术支持。

关键词：剑麻; AsKNOX I; 真核表达; 原核表达

收稿：2015-02-06   修定：2015-03-10

资助：国家现代农业产业技术体系建设专项(CARS-19)。

Cloning and Expression of AsKNOX I of Agave sisalana

LU Jun-Ying, ZHANG Yan-Mei, LI Jun-Feng, ZHOU Wen-Zhao*

Hainan Provincial Key Laboratory for Tropical Crops Nutrition, South Subtropical Crop Research Institute, Chinese Academy of Tropical Agricultural Sciences, Zhanjiang, Guangdong 524091, China

Corresponding author: ZHOU Wen-Zhao; E-mail: wenzhaozhou@163.com; Tel: 0759-2859216

Abstract:

Plant knotted1-like homeobox genes (KNOX) are closely reated with leaf formation. A full-length gene of AsKNOX I, encoding 357 amino acids, was amplified from the shoot apical meristem (SAM) of sisal (Agave sisalana). The predicted molecular weight of AsKNOX I is about 40 kDa. Isoelectric point (pI) is 6.4. The protein includes some conservative domain structures of KNOX I family such as HD, MEINOX, ELK, and GSE. Therefore, the prokaryotic and eukaryotic expression vectors were consctructed, and expressed in Escherichia coli and tobacco leaf, respectively. The AsKNOX 1 protein was purified with His tags. Western blot was used to detect the expression product. The results indicated that AsKNOX 1 had been successful expressed in prokaryotic and eukaryotic expression system, which will provide a technical support for further analyzing the protein structure and function.

Key words: Agave sisalana; AsKNOX I; prokaryotic expression; eukaryotic expression