Plant Cell, Tissue and Organ Culture

, Volume 84, Issue 1, pp 69–79

Agrobacterium-mediated transformation of American chestnut (Castanea dentata (Marsh.) Borkh.) somatic embryos

  • Linda D. Polin
  • Haiying Liang
  • Ronald E. Rothrock
  • Mutsumi Nishii
  • Deborah L. Diehl
  • Andrew E. Newhouse
  • C. Joseph Nairn
  • William A. Powell
  • Charles A. Maynard
Article

DOI: 10.1007/s11240-005-9002-1

Cite this article as:
Polin, L.D., Liang, H., Rothrock, R.E. et al. Plant Cell Tiss Organ Cult (2006) 84: 69. doi:10.1007/s11240-005-9002-1

Abstract

These studies were designed to test if a binary vector containing the gfp, bar and oxalate oxidase genes could transform American chestnut somatic embryos; to see if a desiccation treatment during co-cultivation would affect the transformation frequency of different American chestnut somatic embryo clones; to explore the effects of more rapid desiccation; and to see if the antibiotics used to kill the Agrobacterium were interfering with the regeneration of the somatic embryos. Two days of gradual desiccation was found to significantly enhance transient GFP expression frequency. When this treatment was tested on six American chestnut clones, five were transformed and four of these remained embryogenic. Transformation was confirmed by Southern hybridization. Phenotypically normal transgenic shoots were regenerated and rooted. Vascular tissue specific expression of the oxalate oxidase gene was detected in one transgenic line. Carbenicillin, cefotaxime, and tricarcillin were found to not interfere with the regeneration of transformed embryos.

Key words:

barclonal variationdesiccationgfpoxalate oxidaseregeneration

Abbreviations

2,4-D

2,4-dichlorophenoxyacetic acid

BA

6-benzyladenine

bar

bialaphos-resistance gene

Carb

carbenicillin

Cef

cefotaxime

mgfp5-ER

modified green fluorescent protein gene

OxO

oxalate oxidase

PCR

polymerase chain reaction

PPT

phosphinothricin

SDS

sodium dodecyl sulfate

SSC

sodium chloride sodium citrate

TE

Tris–EDTA

Tric

tricarcillin

WPM

McCown’s woody plant medium salts

Copyright information

© Springer Science+Business Media, Inc. 2005

Authors and Affiliations

  • Linda D. Polin
    • 1
  • Haiying Liang
    • 4
  • Ronald E. Rothrock
    • 1
  • Mutsumi Nishii
    • 2
  • Deborah L. Diehl
    • 1
  • Andrew E. Newhouse
    • 2
  • C. Joseph Nairn
    • 3
  • William A. Powell
    • 2
  • Charles A. Maynard
    • 1
  1. 1.Faculty of Forest and Natural Resources ManagementState University of New York, College of Environmental Science and ForestrySyracuseUSA
  2. 2.Faculty of Environmental and Forest BiologyState University of New York, College of Environmental Science and ForestrySyracuseUSA
  3. 3.Daniel B. Warnell School of Forest ResourcesUniversity of GeorgiaAthensUSA
  4. 4.School of Forest Resources, Department of Horticulture and Huck Institute for Life SciencesPennsylvania State UniversityUniversity ParkUSA