Abstract
Gene transformation is an integral tool for plant genetic engineering. All antibiotic resistant genes currently employed are of bacterial origin and their presence in the field is undesirable. Therefore, we developed a novel and efficient plant native non-antibiotic selection system for the selection of transgenic plants in the model system Arabidopsis. This new system is based on the enhanced expression of Arabidopsis tryptophan synthase beta 1 (AtTSB1) and the use of 5-methyl-tryptophan (5MT, a tryptophan [Trp] analog) and/or CdCl2 as selection agent(s). We successfully integrated an expression cassette containing an AtTSB1 cDNA driven by a cauliflower mosaic virus 35S promoter into Arabidopsis by floral dip transformation. Transgenic plants were efficiently selected on MS medium supplemented with 75 μM 5MT or 300 μM CdCl2 devoid of antibiotics. TSB1 selection was as efficient as the conventional hygromycin selection system. Northern blot analysis of transgenic plants selected by 5MT and CdCl2 revealed increased TSB1 mRNA transcript whereas uneven transcript levels of hygromycin phosphotransferase II (hpt) (control) was observed. Gas chromatography-mass spectrometry revealed 10–15 fold greater free Trp content in AtTSB1 transgenic plants than in wild-type plants grown with or without 5MT or CdCl2. Taken together, the TSB1 system provides a novel selection system distinct from conventional antibiotic selection systems.
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Abbreviations
- TSB1:
-
Tryptophan synthase beta 1 gene
- 5MT:
-
5′ Methyl-tryptophan
- Trp:
-
Tryptophan
- CdCl2 :
-
Cadmium chloride
- TS-α:
-
Tryptophan synthase-α
- TS-β:
-
Tryptophan synthase-β
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Acknowledgments
We are grateful to The Institute of Molecular Biology, Academia Sinica, for providing experimental equipment and facility. This work was supported by a grant from Academia Sinica and grant 94.AS-5.2.1-ST-a1 from the National Science and Technology Program for Agriculture Biotechnology of the Republic of China. Paoyuan Hsiao and Sanjaya contributed equally to this work. We also thank anonymous reviewers for useful comments.
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Hsiao, P., Sanjaya, Su, RC. et al. Plant native tryptophan synthase beta 1 gene is a non-antibiotic selection marker for plant transformation. Planta 225, 897–906 (2007). https://doi.org/10.1007/s00425-006-0405-y
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DOI: https://doi.org/10.1007/s00425-006-0405-y