基于报告基因敲入构建果生刺盘孢细胞核荧光标记菌株  

Fluorescent labeling of Colletotrichum fructicola nuclei based on a reporter gene knock-in strategy

作　　者：董秋月 梁晓飞[1] 张玮[1] 孙广宇[1] 张荣[1] 

机构地区：[1]西北农林科技大学植物保护学院,陕西杨凌712100

出　　处：《菌物学报》2018年第2期166-174,共9页Mycosystema

基　　金：国家重点研发计划(2016YFD0201100);现代农业产业技术体系(CARS-28)~~

摘　　要：运用基因敲入技术,将GFP、mCherry整合到果生刺盘孢Colletotrichum fructicola组蛋白histone H1位点,实现融合表达,获得细胞核荧光标记菌株。基于标记菌株可以对分生孢子、营养菌丝、附着胞、侵染菌丝等结构中的细胞核进行实时活体观察。果生刺盘孢存在性亲和的"+"、"-"型菌株分化,GFP"+"型菌株和m Cherry"-"型菌株接触形成明显杂交线,杂交线上单子囊内含红绿两种孢子,表明"+"、"-"型菌株间发生杂交;杂交线上子囊壳壁表达m Cherry,表明由"-"型菌株发育而来。本研究构建的核荧光标记菌株将是研究果生炭疽菌细胞周期调控和有性繁殖过程的重要材料。Based on a gene knock-in strategy, the reporter gene GFP （or mCherry） was fused with a histone H1 gene and the cassettes were transformed and incorporated into the genome of Colletotrichum fructicola for in-fusion expression and nuclear labeling. Based on the obtained strains, live image of nuclear behavior was possible for various fungal structures such as conidium, vegetative hyphae, appressorium, and infectious hyphae. C. fructicola contains sexually compatible ‘plus’ and ‘minus’ strains, GFP-labeled ‘plus’ strain and mCherry-labeled ‘minus’ strain formed a ‘mating line’ at the contact zone. Both mCherry-labeled and GFP-labeled ascospores could be observed within the same ascus at the ‘mating line’, demonstrating crossing events between the ‘plus’ and ‘minus’ strains. The cell walls of ‘mating line’ perithecia expressed mCherry, demonstrating their ‘minus’ strain derivation. The nuclear-labeled strains reported here allow for convenient and direct observation of nuclear behaviors, and will be important materials for studying cell cycle regulations and sexual reproductions of C. fructicola.

关 键 词：小丛壳属 子囊壳 荧光蛋白 基因敲入 有性繁殖 

分 类 号：S432.44[农业科学—植物病理学]