Transgenic Research

, Volume 16, Issue 6, pp 771–781

pORE: a modular binary vector series suited for both monocot and dicot plant transformation

  • Catherine Coutu
  • James Brandle
  • Dan Brown
  • Kirk Brown
  • Brian Miki
  • John Simmonds
  • Dwayne D. Hegedus
Original Paper

DOI: 10.1007/s11248-007-9066-2

Cite this article as:
Coutu, C., Brandle, J., Brown, D. et al. Transgenic Res (2007) 16: 771. doi:10.1007/s11248-007-9066-2

Abstract

We present a series of 14 binary vectors suitable for Agrobacterium-mediated transformation of dicotyledonous plants and adaptable for biolistic transformation of monocotyledonous plants. The vector size has been minimized by eliminating all non-essential elements from the vector backbone and T-DNA regions while maintaining the ability to replicate independently. The smallest of the vector series is 6.3 kb and possesses an extensive multiple cloning site with 21 unique restriction endonuclease sites that are compatible with common cloning, protein expression, yeast two-hybrid and other binary vectors. The T-DNA region was engineered using a synthetic designer oligonucleotide resulting in an entirely modular system whereby any vector element can be independently exchanged. The high copy number ColE1 origin of replication has been included to enhance plasmid yield in Escherichia coli. FRT recombination sites flank the selectable marker cassette regions and allow for in planta excision by FLP recombinase. The pORE series consists of three basic types; an ‘open’ set for general plant transformation, a ‘reporter’ set for promoter analysis and an ‘expression’ set for constitutive expression of transgenes. The sets comprise various combinations of promoters (PHPL, PENTCUP2 and PTAPADH), selectable markers (nptII and pat) and reporter genes (gusA and smgfp).

Keywords

Agrobacterium tumefaciens Binary vector Plant transformation T-DNA Promoter Selectable marker 

Abbreviations

bp

Base pair

DNA

Deoxyribonucleic acid

dsDNA

Double stranded DNA

gusA

β-Glucuronidase

LB

Left T-DNA border

MCS

Multiple cloning site

nptII

Neomycin phosphotransferase II

OxOx

Oxalate oxidase

pat

Phosphinothricin acetyltransferase

PENTCUP2

Tobacco cryptic constitutive promoter

PCR

Polymerase chain reaction

PHPL

Arabidopsis thaliana hydroperoxide lyase promoter

PTAP

Triticum aestivum lipid transfer protein promoter

PTAPADH

Triticum aestivum lipid transfer protein promoter fused to an alcohol dehydrogenase intron

RB

Right T-DNA border

SDM

Site directed mutagenesis

smgfp

soluble modified green fluorescent protein

T-DNA

Transferred DNA

Ti

Tumour inducing

TNOS

Polyadenylation signal from the nopaline synthase gene

Vir

Virulence region of the Ti plasmid

Copyright information

© Springer Science+Business Media B.V. 2007

Authors and Affiliations

  • Catherine Coutu
    • 1
  • James Brandle
    • 2
  • Dan Brown
    • 2
  • Kirk Brown
    • 2
  • Brian Miki
    • 3
  • John Simmonds
    • 3
  • Dwayne D. Hegedus
    • 1
  1. 1.Agriculture and Agrifood Canada, Saskatoon Research CentreSaskatoonCanada
  2. 2.Agriculture and Agri-Food Canada, Southern Crop Protection and Food Research CenterLondonCanada
  3. 3.Agriculture and Agri-Food Canada, Eastern Cereal and Oilseed Research CentreOttawaCanada

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