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Expression of a Chitinase Gene from Metarhizium anisopliae in Tobacco Plants Confers Resistance against Rhizoctonia solani

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Abstract

The chit1 gene from the entomopathogenic fungus Metarhizium anisopliae, encoding the endochitinase CHIT42, was placed under the control of the CaMV 35S promoter, and the resulting construct was transferred to tobacco. Seventeen kanamycin-resistant transgenic lines were recovered, and the presence of the transgene was confirmed by polymerase chain reactions and Southern blot hybridization. The number of chit1 copies was determined to be varying from one to four. Copy number had observable effects neither on plant growth nor development. Substantial heterogeneity concerning production of the recombinant chitinase, and both general and specific chitinolytic activities were detected in leaf extracts from primary transformants. The highest chitinase activities were found in plants harboring two copies of chit1 inserts at different loci. Progeny derived from self-pollination of the primary transgenics revealed a stable inheritance pattern, with transgene segregation following a mendelian dihybrid ratio. Two selected plants expressing high levels of CHIT42 were consistently resistant to the soilborne pathogen Rhizoctonia solani, suggesting a direct relationship between enzyme activity and reduction of foliar area affected by fungal lesions. To date, this is the first report of resistance to fungal attack in plants mediated by a recombinant chitinase from an entomopathogenic and acaricide fungus.

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Acknowledgements

We thank Dr. A.T.S. Matsumura for providing the isolates of R. solani employed in this study, Dr. M.H. Bodanese-Zanettini for green house facilities, Dr. S.C. Jacques for her assistance with statistical analysis, and architect R. Pasquali for his help with the illustrations. This research was supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq/Brazil) and Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS).

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Authors and Affiliations

  1. Graduating Program in Genetics and Molecular Biology of the Biosciences Institute, Biotechnology Center, Federal University of Rio Grande do Sul (UFRGS), Porto Alegre, RS, Brazil

    Marcelo Fernando Kern, Simone de Faria Maraschin, Débora Vom Endt & Giancarlo Pasquali

  2. State University of Rio Grande do Sul (UERGS), Porto Alegre, RS, Brazil

    Débora Vom Endt

  3. Graduating Program in Cell and Molecular Biology, Biotechnology Center, UFRGS, Porto Alegre, RS, Brazil

    Augusto Schrank, Marilene Henning Vainstein & Giancarlo Pasquali

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  1. Marcelo Fernando Kern
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Correspondence to Giancarlo Pasquali.

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Kern, M.F., Maraschin, S.d.F., Vom Endt, D. et al. Expression of a Chitinase Gene from Metarhizium anisopliae in Tobacco Plants Confers Resistance against Rhizoctonia solani . Appl Biochem Biotechnol 160, 1933–1946 (2010). https://doi.org/10.1007/s12010-009-8701-1

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