Identification Of New Disease Caused By Fusarium Graminearum Diseases And Functional Analysis Of Its Fg2518 Gene

Fusarium graminearum can cause ear rot,stem rot,root rot and other symptoms of wheat and other cereal plants,and eventually cause crop yield reduction.F.graminearum hhwill secrete some secondary metabolites during infection,including mycotoxins harmful to humans and animals.The mycotoxins produced by F.graminearum are stable and can enter the food chain along with food or feed processing and harm human health.Therefore,the prevention and control of F.graminearum and its mycotoxins has always been a hot issue in the prevention and control of crop diseases.The infection of F.graminearum on ryegrass and other herbage was rarely reported.In this study,a new disease of ryegrass caused by Fusarium was found in Hongya County,Sichuan Province.In order to understand the infection and transmission of F.graminearum on herbages and identify the species status of the pathogen,we identified the pathogen.The genomic DNA of isolated strains was amplified by PCR using specific identification primers such as PHO primers and EF primers.Through bioinformatics analysis,the pathogen causing ryegrass disease was finally determined to be Fusarium meridionale.It belongs to Fusarium graminearum species complex(FGSC).Further experiments showed that the radial growth rate of F.meridionale was about 11.0 mm.The mespore was sickle-shaped to nearly straight,with 3-5 intervals.Fg2518 gene is a gene that may be related to circular RNA discovered by transcriptome sequencing in our laboratory.In this experiment,Fg2518 gene knockout fragments were constructed through overlap,and then the knockout fragments were introduced into the protoplast of PH-1 of F.graminearum wild-type strain by PEG mediated method.Through the principle of homologous recombination,Fg2518 gene was replaced by screening marker genes and Fg2518 gene was knocked out.HYG antibiotics were used for preliminary screening of translators,and multiple pairs of specific primers such as 2518-JD and H853 were used to identify whether the single colony was a positive mutant.Finally,two complement mutants were constructed based on Fg2518 positive mutants using the same principle and method.One was a complete callback mutant of Fg2518 gene,and the other was a callback mutant with deletion of the circular RNA encoding part of Fg2518 gene.Finally,the obtained Fg2518 generelated mutant was used together with the wild-type strain to study the strain phenotype,such as the growth rate,stress resistance,sporulation ability,pathogenicity and virulence ability of the strain.In the growth rate assay experiment,the growth rate of Fg2518 gene deletion mutant was significantly lower than that of the wild type strain,so it can be speculated that Fg2518 gene is related to the growth regulation of F.graminearum.In the stress experiment,there was no significant change in the growth rate among the mutant strains,so it can be inferred that Fg2518 deletion had no effect on the stress resistance of F.graminearum.The sporulation capacity of Fg2518 mutant was significantly increased compared with the wild-type strain PH-1,while the spore length was significantly decreased,and the spore width was not significantly changed.Therefore,Fg2518 gene may be related to the sporulation ability of F.graminearum.In the coleoptile experiment,rice toxigenesis experiment and TBI induction culture medium toxigenesis experiment,it was found that the pathogenicity and virulence ability of Fg2518 gene deletion mutant were not significantly changed compared with the wild type strain,but the complement mutant with deletion of the gene fragment encoding circular RNA in Fg2518 gene was significantly reduced in both pathogenicity and virulence ability.So the gene fragment that encodes circular RNA or this circular RNA has a regulatory role in the biosynthesis of mycotoxins such as DON toxin in F.graminearum.Moreover,the non-coding parts of Fg2518 gene may have regulatory elements that negatively regulate the formation of circular RNA.In this study,the biological function of Fg2518 gene was investigated.The experimental results showed that Fg2518 gene plays a regulatory role in the growth and reproduction of F.graminearum and the synthesis of secondary metabolites such as DON toxin.This laid a foundation for the subsequent experimental study on the regulatory pathways of Fg2518 gene regulating mycotoxins such as DON toxin,and also provided a new research angle for the control of F.graminearum.