Transformation of Botrytis cinerea with a Green Fluorescent Protein (GFP) Gene for the Study of Host-pathogen Interactions

摘要

Botrytis cinerea is a plant pathogenic fungus that causes diseases generally known as gray mold in a wide variety of agriculturally important crops. To effectively reduce gray mold diseases and minimize synthetic chemical use on fruits pre- and postharvest, it is necessary to have an understanding on latent infections and the behaviours of the pathogen under various treatments. We described here a procedure for B. cinerea transformation based on particle bombardment. Utilising a co-trartsformation system, we successfully introduced a Green Fluorescent Protein (GFP) reporter gene and a hygromycin B resistant (Hy*) selectable marker into the fungal conidia. Within the five Hy positive colonies, one isolate BC-2b that displayed strong green fluorescence under a fluorescent compound microscope confirmed the GFP gene insertion by direct PCR. Morphological observation, cultural evaluation and pathogenicity tests on flower petals and fruits of strawberry revealed that the GFP transformant of B, cinerea maintained the characteristics of the wild type isolate and was able to express the GFP gene in hyphae and conidia of the fungus both in vitro and in vivo. This proved that the transformant could be a powerful tool for our future studies on the interactions between the pathogen and its fruit hosts.