Broad-specificity amino acid racemase, a novel non-antibiotic selectable marker for transgenic plants

  • Yi-Chia Kuan
  • Venkatesan Thiruvengadam
  • Jia-Shin Lin
  • Jia-Hsin Liu
  • Tsan-Jan Chen
  • Hsin-Mao Wu
  • Wen-Ching Wang
  • Liang-Jwu Chen
Original Article
  • 47 Downloads

Abstract

The broad-specificity amino acid racemase (Bsar) from Pseudomonas putida catalyzes the racemization of various amino acids, offering a flexible and feasible platform to develop a new non-antibiotic selectable marker system for plant transformation. In the present study, we demonstrated that a Bsar variant, Bsar-R174K, that is useful as a selectable marker gene in Arabidopsis and rice that were susceptible to l-lysine and D-alanine. The introduction of wild-type Bsar, Bsar-R174K or Bsar-R174A into E. coli lysine or asparagine auxotrophs was able to rescue the growth of these microorganisms in minimal media supplemented with selectable amino acid enantiomers. The transformation of Arabidopsis with Bsar or Bsar variants based on d-alanine selection revealed that Bsar-R174K had the greatest efficiency (2.40%), superior to kanamycin selection-based transformation (1.10%). Whereas, l-lysine-based selection exhibited lower efficiency for Bsar-R174K (0.17%). The progenies of selected Bsar-R174K transgenic Arabidopsis revealed normal growth properties. In addition, Bsar-R174K transgenic rice was obtained on l-lysine medium with an efficiency of 0.9%, and the progenies of the transgenic rice revealed morphologically normal phenotypes comparable with their wild-type counterparts. This study presents the first report of broad range amino acid racemase Bsar-R174K as a non-antibiotic selectable marker system applied in transgenic plants.

Keywords

Plant transformation Broad-specificity amino acid racemase Non-antibiotic marker Transgenic rice 

Notes

Acknowledgements

This work was financially supported through grants from the Ministry of Science and Technology, Taiwan (101-2321-B-007-004, 102-2321-B-007-003, and 103-2321-B-007 -002).

Compliance with ethical standards

Conflict of interest

The authors declare no competing financial interests.

Supplementary material

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Supplementary material 1 (DOCX 13 KB)
11816_2018_469_MOESM2_ESM.pptx (9.2 mb)
Supplementary material 2 (PPTX 9462 KB)

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Copyright information

© Korean Society for Plant Biotechnology and Springer Japan KK, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Institute of Molecular and Cellular Biology and Department of Life SciencesNational Tsing-Hua UniversityHsinchuTaiwan
  2. 2.Institute of Molecular BiologyNational Chung Hsing UniversityTaichungTaiwan
  3. 3.Department of BiotechnologyHungkuang UniversityTaichungTaiwan
  4. 4.Department of Plant Genetic Resources, Centre for Plant Breeding and GeneticsTamil Nadu Agricultural UniversityCoimbatoreIndia
  5. 5.Agricultural Biotechnology CenterNational Chung Hsing UniversityTaichungTaiwan

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