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Gene silencing assessment for genes from recalcitrant or poorly studied plant species

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Abstract

We have developed an efficient system of assessing the ability of a gene silencing cassette to silence transcripts from recalcitrant or poorly studied plant species by using a model plant as a host for the gene of interest. Tobacco plants transgenic for Lachrymatory Factor Synthase (LFS) enzyme activity from onion were first produced by introducing a CaMV 35S-onion-lfs gene construct. These plants were then subjected to a second transformation with an RNAi construct directed against the lfs gene sequence. LFS enzyme activity assay showed that the transgenic plants, containing both the lfs gene and the RNAi construct, had significantly reduced LFS activity. This observation was supported by Western analysis for the LFS protein and further validated by quantitative RT-PCR analysis that demonstrated a significant reduction in the lfs transcript level in the dual transformants. In this work, we have demonstrated that the RNAi construct is a suitable candidate for the development of a non-lachrymatory onion. Our model plant RNAi system has wide-reaching applications for assessment and targeting of plant secondary pathway genes, from poorly studied or recalcitrant plant species, that are important in the pharmacological, food and process industries.

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Acknowledgments

We thank CSIRO Plant Industry, Canberra, Australia, for use of RNAi technology. We are grateful to Sheree Davis (Crop & Food Research) for maintenance of tobacco plants. Finally, special thanks to Prof. Tony Conner (Crop & Food Research, Lincoln University) and Dr. Chris Winefield (Lincoln University) for their advice during this study.

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Authors and Affiliations

  1. Somatech Center, House Foods Corporation, 1-4 Takanodai, Yotsukaido, Chiba, 284-0033, Japan

    Takahiro Kamoi & Shinsuke Imai

  2. New Zealand Institute for Crop and Food Research Limited, Private Bag 4704, Christchurch, New Zealand

    Colin Charles Eady

  3. National Centre for Advanced Bio-Protection Technologies, Lincoln University, P.O. Box 84, Canterbury, New Zealand

    Takahiro Kamoi & Colin Charles Eady

Authors
  1. Takahiro Kamoi
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  2. Colin Charles Eady
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  3. Shinsuke Imai
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Correspondence to Takahiro Kamoi.

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Kamoi, T., Eady, C.C. & Imai, S. Gene silencing assessment for genes from recalcitrant or poorly studied plant species. Plant Biotechnol Rep 2, 199–206 (2008). https://doi.org/10.1007/s11816-008-0062-7

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  • DOI: https://doi.org/10.1007/s11816-008-0062-7

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