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RNA Interference-Based Transgenic Maize Resistant to Maize Dwarf Mosaic Virus

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Abstract

Maize dwarf mosaic virus (MDMV) is a widespread pathogenic virus that causes serious loss of yield in maize (Zea mays). RNA interference (RNAi) triggered by hairpin RNA (hpRNA) transcribed from a transgenic inverted-repeat sequence is an effective way to defend against viruses in plants. In this study, an hpRNA expression vector containing a sense arm and an antisense arm of 150 bp separated by an intron of the maize actin gene was constructed to target the P1 protein (protease) gene of MDMV and used to transform Agrobacterium tumefaciens strain EHA105. The transformed Agrobacterium strain was used to transform maize embryonic calli isolated from immature embryos by an improved culture technique. In all, 46 plants were regenerated after stringent hygromycin B selection, and 18 of them were certified to be positive by PCR amplification. Of these positive plants, 13 were grown to produce offspring, and nine were identified by Southern blotting to have the transgene integrated with one or two copies. The resistance of three T2 lines was evaluated in a field trial of dual MDMV inoculation in two environments and was found to be improved compared with the non-transformed control. The disease indexes of the transgenic plant lines h2, 13, and h1 were not significantly different from the highly resistant control line H9-21. The viral titers of the inoculated plants were detected by double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), and the result was in accord with the resistance evaluated in the field trial. The addition of uniconazole S3307 (0.25 mg l−1) and ABT root-promoting powder (0.5 mg l−1) showed a significant improvement of hardening in regenerated plantlets, which were stronger and generated a better fibrous root system than the control. This improvement could facilitate the transgenic operation of maize.

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Abbreviations

IgG:

Immunoglobulin G

T-DNA:

Transfer DNA

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Acknowledgements

Financial support from the National Key Science and Technology Special Project (2008ZX08003-004 and 2009ZX08003-012B) and technical support from all the staff members to this project are sincerely appreciated. The English writing has been revised by the International Science Editing.

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Authors and Affiliations

  1. Maize Research Institute, Sichuan Agricultural University, Xingkang Road 46, 625014, Ya’an, Sichuan, China

    Zhi-Yong Zhang, Feng-Ling Fu & Wan-Chen Li

  2. Faculty of Biology and Science, Sichuan Agricultural University, Xingkang Road 36, 625014, Ya’an, Sichuan, China

    Lin Gou & Han-Guang Wang

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  1. Zhi-Yong Zhang
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  2. Feng-Ling Fu
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Correspondence to Wan-Chen Li.

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Zhang, ZY., Fu, FL., Gou, L. et al. RNA Interference-Based Transgenic Maize Resistant to Maize Dwarf Mosaic Virus. J. Plant Biol. 53, 297–305 (2010). https://doi.org/10.1007/s12374-010-9117-8

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