小麦赤霉菌FGSG__04871基因敲除及功能研究

Research on Knockout and Function of FGSG__04871 Gene in Fusarium graminearum

应用Split-Marker基因敲除技术,构建含有潮霉素抗性基因hph的基因敲除盒,由PEG介导,进行原生质体转化,在含有潮霉素的培养基上筛选转化子,应用PCR正负筛查确定缺失突变体.根据缺失突变体致病性的检测及表型变化对FGSG_04871基因的生物学功能进行分析.通过PCR正负筛选成功获得了3个敲除突变体,分别命名为△FGSG_04871 1-1、△FGSG_04871 1-3、△FGSG_04871 2-1.以野生型PH-1作为对照,敲除突变体的菌落形态没有明显变化,菌落生长速度略微滞后,致病力没有减弱.但是,突变体的产孢量低于野生型PH-1,因此,FGSG_04871基因可能与小麦赤霉菌分生孢子的生长能力有关.

The Split-marker method was applied to construct the deletion cassette of the FGSG_04871 gene. The primers were designed according the sequence in the Fusarium Genome Database and the PCR products were transformed into the protoplast of wild type PH-1 by the PEG-mediated transformation method. Transformants were selected through medium containing hygromycin and then the knockout mutants were confirmed by PCR using positive and negative primers. Three deletion mutant of FGSG_04871 was obtained in this research, named △FGSG_04871 1-1、△FGSG_04871 1-3 and △FGSG_04871 2-1 respectively. Compared with the wild type, the growth rate of FGSG_04871 deletion mutant was small slightly. There was no significant difference between mutant and wild type PH-1 in colo- ny morphology and conidium morphology. The result of tomato infection experiment showed that deletion of FGSG_04871 gene did not reduce in virulence. However,the spore amount of deletion mutant was less than PH-1 ,therefore, FGSG_04871 gene may be related to conidia formation in Fusariurn graminearum.