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A bacterial gene codA encoding cytosine deaminase is an effective conditional negative selectable marker in Glycine max

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Abstract

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Research describes the practical application of the codA negative selection marker in Soybean. Conditions are given for codA selection at both the shooting and rooting stages of regeneration.

Abstract

Conditional negative selection is a powerful technique whereby the absence of a gene product allows survival in otherwise lethal conditions. In plants, the Escherichia coli gene codA has been employed as a negative selection marker. Our research demonstrates that codA can be used as a negative selection marker in soybean, Glycine max. Like most plants, soybean does not contain cytosine deaminase activity and we show here that wild-type seedlings are not affected by inclusion of 5-FC in growth media. In contrast, transgenic G. max plants expressing codA and grown in the presence of more than 200 μg/mL 5-FC exhibit reductions in hypocotyl and taproot lengths, and severe suppression of lateral root development. We also demonstrate a novel negative selection-rooting assay in which codA-expressing aerial tissues or shoot cuttings are inhibited for root formation in media containing 5-FC. Taken together these techniques allow screening during either the regeneration or rooting phase of tissue culture.

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Acknowledgments

We are grateful to Ron Chan, Bryan Hernandez, and Paul Duellman for technical assistance. This work was supported by the United Soybean Board project number 1420-532-5644, by the Minnesota Soybean Research and Promotion Council project number 7-14C, by the USDA Agricultural Research Service CRIS project 5325-21000-020, and by the Biotechnology Risk Assessment Program competitive grant 2010-33522-21773 from the USDA—National Institute of Food and Agriculture. Mention of trade names or commercial products is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the US Department of Agriculture. USDA is an equal opportunity provider and employer.

Conflict of interest

The authors declare they have no conflict of interest.

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Authors and Affiliations

  1. Department of Plant Sciences, University of California-Davis, Davis, CA, 95616, USA

    Min Shao & Sara K. Hotton

  2. Department of Agronomy and Plant Genetics, University of Minnesota, St. Paul, MN, 55108, USA

    Jean-Michel Michno

  3. Western Regional Research Center, USDA-ARS Crop Improvement and Genetics, Albany, CA, 94710, USA

    Ann Blechl & James Thomson

Authors
  1. Min Shao
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  2. Jean-Michel Michno
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  3. Sara K. Hotton
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  4. Ann Blechl
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  5. James Thomson
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Corresponding author

Correspondence to James Thomson.

Additional information

Communicated by K. Wang.

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Shao, M., Michno, JM., Hotton, S.K. et al. A bacterial gene codA encoding cytosine deaminase is an effective conditional negative selectable marker in Glycine max . Plant Cell Rep 34, 1707–1716 (2015). https://doi.org/10.1007/s00299-015-1818-5

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  • DOI: https://doi.org/10.1007/s00299-015-1818-5

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