Identification PG Gene Family Of Cuvrularia Lunata And The Role Of Clpg1 In Pathogenecity

Curvularia leaf spot of maize, caused by Curvularia lunata(Wakker) Boed, is one of the most important diseases in northern China’s maize production zone, and is currently one of the most widely distributed maize leaf diseases nationwide. In this study, the polygalacturonase gene family of Curvularia lunata were identified and characterized by bioinformatics, and the expression pattern in pathogen-plant interaction and in different growth stages under pectin-induced were analyzed. We had identified the role of Clpg1 in pathogenecity by gene knockout technology. The main results are as follows:1. We identified four members from polygalacturonase gene family in Curvularia lunata based on bioinformatics, namely Clpg1、Clpg2、Clpg3 and Clpg4. In the four proteins, the longest amino acid sequence containing 449 amino acid residues, the shortest containing 273 amino acid residues, protein molecular weight in the range of 27.4 ~ 48.5 kD, isoelectric point in the range of 5.53 ~ 9.19. Through the analysis of the structure and conserved domain of the Clpg,we found all the genes have same exon and intron numbers and the same conserved domain with other fungi. The Clpg1, Clpg2, Clpg3 and Clpg4 is divided into two major branches using phylogenetic analysis. The Clpg1, Clpg2 and Clpg3 were in the same branch, showing a close genetic relationship with Bipolaris oryzae, Stemphylium lycopersici, Pyrenophora tritici-repentis and Setosphaeria turcica, Clpg3 has a close genetic relationship with Bipolaris maydis in the same branch.2. The expression pattern of PG genes in C. lunata were analyzed during different growth stage under pectin-induced and pathogen-host interactions periods by real-time PCR. The results showed that Clpg1, Clpg2 and Clpg4 have highest expression at 2 d, Clpg3 have highest expression at 4 in different growth stage under pectin-induced. In pathogen-host interactions,Clpg1 and Clpg3 were reached the highest at 3 h inoculation, and the expression of Clpg2 and Clpg4 gradually increased, peaked at 24 h inoculation. These results indicated that PG genes may play an important role in pathogenecity.3. Clpg1 knockout vector were constructed by double-joint PCR. In total, 79 resistance transformants were obtained by PEG-mediated genetic transformation. One of those were identified as Clpg1 gene knockout mutant transformant(△Clpg1).4. The biological characteristics and pathogenecity of △Clpg1 were analyzed. The results showed that the conidia germination rate and conidia production of △Clpg1 were decreased and the polygalacturonase activity of △Clpg1 reduced by 16% compared to wild-type strain DQ-1.In pathogenecity experiments, we found that the pathogenecity of △ Clpg1 were decreased compared to wild-type strain DQ-1. The results indicated that Clpg1 may influence polygalacturonase activity, resulting in the decrease of Curvularia lunata pathogenecity.