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Retransformation of Marker-Free Potato for Enhanced Resistance Against Fungal Pathogens by Pyramiding Chitinase and Wasabi Defensin Genes

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Abstract

Multi-auto-transformation vector system has been one of the strategies to produce marker-free transgenic plants without using selective chemicals and plant growth regulators and thus facilitating transgene stacking. In the study reported here, retransformation was carried out in marker-free transgenic potato CV. May Queen containing ChiC gene (isolated from Streptomyces griseus strain HUT 6037) with wasabi defensin (WD) gene (isolated from Wasabia japonica) to pyramid the two disease resistant genes. Molecular analyses of the developed shoots confirmed the existence of both the genes of interest (ChiC and WD) in transgenic plants. Co-expression of the genes was confirmed by RT-PCR, northern blot, and western blot analyses. Disease resistance assay of in vitro plants showed that the transgenic lines co-expressing both the ChiC and WD genes had higher resistance against the fungal pathogens, Fusarium oxysporum (Fusarium wilt) and Alternaria solani (early blight) compared to the non-transformed control and the transgenic lines expressing either of the ChiC or WD genes. The disease resistance potential of the transgenic plants could be increased by transgene stacking or multiple transformations.

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Acknowledgments

We are very grateful to Pulp and Paper Research group, Nippon Paper Industries, Tokyo, who kindly provided the MAT vector constructs.

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Authors and Affiliations

  1. Laboratory of Plant Cell Technology, Graduate School of Horticulture, Chiba University, 648 Matsudo, Matsudo, Chiba, 271-8510, Japan

    Raham Sher Khan, Nader Ahmed Darwish, Bushra Khattak, Valentine Otang Ntui, Kynet Kong, Kazuki Shimomae, Ikuo Nakamura & Masahiro Mii

  2. Department of Biotechnology, Abdul Wali Khan University, Mardan, Pakistan

    Raham Sher Khan

  3. Faculty of Agriculture, Aflaton Street, Alshatby-Alexandria, Egypt

    Nader Ahmed Darwish

  4. Department of Genetics and Biotechnology, Faculty of Science, University of Calabar, PMB 1115, Calabar, Nigeria

    Valentine Otang Ntui

  5. Institute of Biotechnology and Genetic Engineering, The University of Agriculture, Peshawar, Pakistan

    Bushra Khattak

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  1. Raham Sher Khan
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Correspondence to Raham Sher Khan.

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Khan, R.S., Darwish, N.A., Khattak, B. et al. Retransformation of Marker-Free Potato for Enhanced Resistance Against Fungal Pathogens by Pyramiding Chitinase and Wasabi Defensin Genes. Mol Biotechnol 56, 814–823 (2014). https://doi.org/10.1007/s12033-014-9760-2

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