title: Transformation of the endochitinase gene Chi67-1 in Clonostachys rosea 67-1 increases its biocontrol activity against Sclerotinia sclerotiorum.
abstract: Clonostachys rosea is a promising biocontrol fungus active against various plant fungal pathogens. In this study, the endochitinase-encoding gene Chi67-1, the expression of which is sharply upregulated in C. rosea 67-1 when induced by sclerotia, was transformed into the original isolate by protoplast transformation, and transformants were screened against Sclerotinia rot of soybean. The transformation efficiency was approximately 50 transformants per 1 × 107protoplasts, and 68 stably heritable recombinants were assayed. The parasitic rates of 32.4% of the tested strains increased by more than 50% compared to 43.3% of the wild type strain in 16 h, and the Rc4-4 transformant showed a parasitic rate of 100% in 16 h. The control efficiencies of the selected efficient transformants to soybean Sclerotinia stem rot were evaluated in pots in the greenhouse, and the results revealed that Rc4-4 achieved the highest efficiency of 81.4%, which was 31.7% and 28.7% higher than the control achieved by the wide type and the pesticide carbendazim, respectively. Furthermore, the expression level of Chi67-1 was 107-fold higher in Rc4-4 than in the wild type, and accordingly, the chitinase activity of the recombinant increased by 140%. The results lay a foundation for the development of efficient genetically engineered strains of C. rosea.
authors.affiliation: Institute of Plant Protection, Chinese Academy of Agricultural Sciences, No. 2 West Yuanmingyuan Road, Haidian District, Beijing, 100193, China.
authors.forename: Zhan-Bin
authors.initials: ZB
authors.lastname: Sun
authors.affiliation: Institute of Plant Protection, Chinese Academy of Agricultural Sciences, No. 2 West Yuanmingyuan Road, Haidian District, Beijing, 100193, China.
authors.forename: Man-Hong
authors.initials: MH
authors.lastname: Sun
authors.affiliation: Institute of Plant Protection, Chinese Academy of Agricultural Sciences, No. 2 West Yuanmingyuan Road, Haidian District, Beijing, 100193, China.
authors.forename: Mo
authors.initials: M
authors.lastname: Zhou
authors.affiliation: Institute of Plant Protection, Chinese Academy of Agricultural Sciences, No. 2 West Yuanmingyuan Road, Haidian District, Beijing, 100193, China. sdli@ippcaas.cn.
authors.forename: Shi-Dong
authors.initials: SD
authors.lastname: Li
keywords: Biocontrol agent; Clonostachys rosea; Endochitinase; Mycoparasitism; Protoplast transformation; Sclerotinia sclerotiorum