甘蓝型油菜MAPK7基因家族及其启动子的克隆与表达分析

从甘蓝型油菜中克隆了MAPK7基因家族3个成员BnMAPK7-1、BnMAPK7-2和BnMAPK7-3的全长cDNA序列，它们最长标准mRNA分别为1593、1434和1547 bp。系统进化分析表明，甘蓝型油菜MAPK7基因全部来自白菜，而且BnMAPK7-1/-2的是由白菜Bra03723祖先基因加倍产生的，而BnMAPK7-3则由白菜中Bra037234的祖先基因进化形成的。同时，克隆获得BnMAPK7-1/-2的启动子序列，该启动子里含有多个与光诱导相关的元件、激素响应元件和逆境胁迫响应元件。实时荧光定量PCR (qRT-PCR)显示甘蓝型油菜MAPK7基因家族在所有的组织器官中均表达，并发现该基因家族的表达受植物激素(MeJA、ABA、SA)、信号分子(H₂O₂)、逆境(高温)以及伤害(损伤和核盘菌)的诱导，初步证明甘蓝型油菜MAPK7基因在植物逆境胁迫应答中起一定作用。实现了甘蓝型油菜MAPK7-1/-3基因的原核表达，并证明它们编码的蛋白主要以包涵体形式存在，为后续的研究提供了基础。

Cloning and Analysis of MAPK7 Gene Family and Their Promoters from Brassica napus

Mitogen-activated protein kinase (MAPK) is a large family of serine/threonine protein kinase, containing 11 conserved subdomains. MAPK pathway play a central role in transferring information from diverse receptors/sensors to a wide range of cellular responses in plants. Signaling through MAP kinase cascade can lead to cellular responses including cell division, development, hormone, physiology, as well as the response to a broad variety of biotic and abiotic stresses. In this study, the full-length cDNA of MAPK7 gene family was isolated from Brassica napus. There were three members in this gene family, including BnMAPK7-1, BnMAPK7-2, and BnMAPK7-3. Standard mRNA lengths of these genes were 1 593, 1 434 and 1 547 bp. Phylogenetic tree showed that BnMAPK7 gene family was all originated from Brassica rapa, and BnMAPK7-1/-2 were generated from ancestral gene of Bra03723, while BnMAPK7-3 was evolved from ancestral gene of Bra03724. We also cloned promoters of BnMAPK7-1/-2, which contain some light responsive elements, hormone responsive elements and stress response elements. Quantitative real-time PCR (qRT-PCR) showed that BnMAPK7 gene family expressed in all organs tested, and could be induced by phytohormones (MeJA, ABA, and SA), signaling molecules (H₂O₂), stress (heat) and injury (wounding and Sclerotinia sclerotiorum). This study preliminarily proved that the BnMAPK7 gene family plays certain roles in plant stress responses. In addition, the recombinant BnMAPK7-1/-3 proteins were successfully expressed in Escherichia coli, but the main form of proteins was inclusion body, this result would contribute to further discuss in the future.