Plant Cell Reports

, Volume 22, Issue 2, pp 122–128

The use of the PMI/mannose selection system to recover transgenic sweet orange plants (Citrus sinensis L. Osbeck)

  • R. L. Boscariol
  • W. A. B. Almeida
  • M. T. V. C. Derbyshire
  • F. A. A. Mourão Filho
  • B. M. J. Mendes
Genetic Transformation and Hybridization

DOI: 10.1007/s00299-003-0654-1

Cite this article as:
Boscariol, R.L., Almeida, W.A.B., Derbyshire, M.T.V.C. et al. Plant Cell Rep (2003) 22: 122. doi:10.1007/s00299-003-0654-1

Abstract

A new method for obtaining transgenic sweet orange plants was developed in which positive selection (Positech) based on the Escherichia coli phosphomannose-isomerase (PMI) gene as the selectable marker gene and mannose as the selective agent was used. Epicotyl segments from in vitro-germinated plants of Valencia, Hamlin, Natal and Pera sweet oranges were inoculated with Agrobacterium tumefaciens EHA101-pNOV2116 and subsequently selected on medium supplemented with different concentrations of mannose or with a combination of mannose and sucrose as a carbon source. Genetic transformation was confirmed by PCR and Southern blot. The transgene expression was evaluated using a chlorophenol red assay and isoenzymes. The transformation efficiency rate ranged from 3% to 23.8%, depending on cultivar. This system provides an efficient manner for selecting transgenic sweet orange plants without using antibiotics or herbicides.

Keywords

Agrobacterium tumefaciensCitrus transformationSweet orangeMannose selection marker

Abbreviations

BAP

Benzylaminopurine

CPR

Chlorophenol red

EGTA

Ethylene glycol-0-0′- bis (2, aminoethyl) N′, N′, N′, N′ tetraacetic acid

MTT

[3-(4,5-Dimethyl thiazol-2-YL)-2,5-diphenyl] tetrazolium bromide

PMI

Phosphomannose isomerase (EC 5.3.1.8)

PMS

Phenazine methosulphate

Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  • R. L. Boscariol
    • 1
  • W. A. B. Almeida
    • 3
  • M. T. V. C. Derbyshire
    • 2
  • F. A. A. Mourão Filho
    • 3
  • B. M. J. Mendes
    • 1
  1. 1.Laboratório de Biotecnologia Vegetal, Centro de Energia Nuclear na AgriculturaUniversidade de São PauloSão PauloBrazil
  2. 2.Laboratório de Melhoramento de Plantas, Centro de Energia Nuclear na AgriculturaUniversidade de São PauloSão PauloBrazil
  3. 3.Departamento de Produção Vegetal, Escola Superior de Agricultura "Luiz de Queiroz"Universidade de São PauloSão PauloBrazil