Plant Cell Reports

, Volume 34, Issue 10, pp 1707–1716 | Cite as

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

  • Min Shao
  • Jean-Michel Michno
  • Sara K. Hotton
  • Ann Blechl
  • James Thomson
Original Paper
First Online:
Received:
Revised:
Accepted:

Abstract

Key message

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.

Keywords

codA gene Conditional negative selection Cytosine deaminase 5-Fluorocytosine Glycine max Soybean 
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Notes

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.

Supplementary material

299_2015_1818_MOESM1_ESM.pdf (272 kb)
Supplementary material 1 (PDF 271 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg (outside the USA) 2015

Authors and Affiliations

  • Min Shao
    • 1
  • Jean-Michel Michno
    • 2
  • Sara K. Hotton
    • 1
  • Ann Blechl
    • 3
  • James Thomson
    • 3
  1. 1.Department of Plant SciencesUniversity of California-DavisDavisUSA
  2. 2.Department of Agronomy and Plant GeneticsUniversity of MinnesotaSt. PaulUSA
  3. 3.Western Regional Research CenterUSDA-ARS Crop Improvement and GeneticsAlbanyUSA

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