Cloning And Functional Analysis Of Antifungal Substance Associated Genes Of Coniothyrium Minitans

Coniothyrium minitans, an important sclerotial mycoparasite of Sclerotinia sclerotiorum, can produce antifungal substances (Antifungal substance, AFS) macrosphelide A in acidic environment, which can inhibit the mycelial growth of Sclerotinia sclerotiorum. In this study, two mutants ZS-1T20440 and ZS-1T8512 obtained from T-DNA insertional library of C. minitans ZS-1 were studied. Compared with the wild strain ZS-1, ZS-1T20440 and ZS-1T8512 could produce more antifungal substances and required a wider range of pH value circumstance. However, the growth rate of them was depressed and they lost the ability to form conidia.Southern blot showed that there was only one copy of T-DNA insertion in ZS-1T20440 and ZS-1T8512. Two AFS-associated genes were clonied and analyzed by TAIL-PCR and RT-PCR. The deduced amino acid sequence cloned from ZS-1T20440 shared high identity with GPI-anchored protein of Aspergillus fumigatus Af293. Therefore the gene cloned here was named CMGPI-like1. The deduced amino acid sequence cloned from ZS-1T8512 included a conserved domain as acyltransferase, therefore the gene was called for CMAT-1. Based on RNAi technique, two vectors were constructed specific to CMGPI-like1 and CMAT-1 and were used to transform ZS-1.7 and 6 transformants were obtained respectively. Further results showed that the ability to form anti-fungal substance of these transformants were increased by diferent extend. Therefore, gene CMGPI-like1 and gene CMAT-1 were related to anti-fungal substances of C. minitans.In addition, we also found some transformants showed few differences in growth and sporulation with ZS-1, but the ability of producing anti-fungal substances was enhanced more than ZS-1. Currently these transformants still need to add further evidences related to molecular biology and so on. The transformants'potential biocontrol research is also in progress.