黑松银松素合成酶的cDNA克隆及表达研究  

Cloning of the cDNA Encoding Pinosylvin Synthase from Pinus Thunbergii and Its Expression in Escherichia Coli

作　　者：杨晓[1] 宋修明 王艳士 李荣贵[1] 

机构地区：[1]青岛大学生命科学学院,山东青岛266071 [2]青岛市森林病虫害防治工作站,山东青岛266000

出　　处：《青岛大学学报（工程技术版）》2014年第2期89-93,共5页Journal of Qingdao University(Engineering & Technology Edition)

基　　金：国家自然科学基金项目(31070575);青岛市科技发展计划项目(12-1-4-2-(1)-jch;12-1-3-46-nsh)

摘　　要：松萎蔫病是由松材线虫引起的一种危害性极大的森林病害,为研究黑松对松材线虫的抗性机制,本文利用RT-PCR技术对黑松体内经转录组分析获得的与银松素合成酶mRNA高相似度的序列进行扩增,获得了银松素合成酶cDNA,然后将此银松素合成酶基因(PSL)克隆到表达载体pET-15b上,构建了重组表达质粒pET-15b-PSL,将其转化大肠杆菌BL21(DE3)后构建工程菌,通过IPTG诱导,工程菌高效表达重组蛋白,经Ni 2+螯合柱亲和层析得到了纯化重组银松素合成酶。该研究为了解黑松银松素合成酶的功能及松萎蔫病抗性松树的培育奠定了基础。Pine wilt disease （PWD） caused by pine wood nematode （Bursaphelenchus xylophilus） is seri- ous thread to forests. In order to study the resistant mechanism of Japanese black pine to pine wood nema- tode, the cDNA coding pinosylvin synthase was amplified by RT-PCR based on the transcriptics data col- lected from Japanese black pine. The cDNA was cloned into pET-15b to construct expressing vector pET- 15b-PLS, and then pET-15b-PLS was transformed into E. coli BL21（DE3） to construct engineering bacte- ria. Recombinant pinosylvin synthase was overexpressed in engineering bacteria by isopropyl [3-D-l-thioga- lactopyranoside （IPTG） induction, and purified by affinity chromatography on a nickel column.. The study will lay a good foundation for further elucidating the functions of pinosylvin synthase and culturing of PWD resistant pines.

关 键 词：黑松 银松素合成酶 CDNA 克隆 表达 

分 类 号：S763.7[农业科学—森林保护学]