Articles
GREEN FLUORESCENT PROTEIN (GFP): A TOOL FOR SELECTING TRANSGENIC PETUNIA PLANTS WITH HIGHER CONSUMER ACCEPTANCE
Article number
953_30
Pages
217 – 222
Language
English
Abstract
The Green Fluorescent Protein (GFP) from the jellyfish Aequorea Victoria has proven to be a convenient and powerful vital marker in transgenic studies.
Its expression can be detected non-destructively, in real time, simply by UV-light excitation.
This property of GFP holds promise in monitoring the presence and expression of transgenes in higher plants.
Based on a transient gene expression assay, the function of two GFP genes was tested: mGFP-4 (Haseloff et al., 1997) and smRS-GFP (Davis and Vierstra, 1998). The intensity of smRS-GFP fluorescence was higher than that of mGFP-4 in Petunia and easier to distinguish from other unspecific fluorescent signals.
First stable transformation experiments in four commercially relevant Petunia cultivars involved the vector pMen65smRS-GFP containing nptII gene.
This vector resulted in clearly detectable GFP expressing callus and made it possible to visually select exclusively for GFP. Selection using GFP only versus GFP and kanamycin selection was compared regarding the transformation efficiencies.
The detection of integration the transgene via southern hybridization revealed single and multiple integrations of smRS-GFP in Petunia. Single copy plants showed intensive expression in all parts of the plants, while rising copy numbers led to only weak or partial expression of smRS-GFP. In this study, the possibility to select transgenic plants based on their GFP expression without applying antibiotic or herbicide resistance genes is shown.
Its expression can be detected non-destructively, in real time, simply by UV-light excitation.
This property of GFP holds promise in monitoring the presence and expression of transgenes in higher plants.
Based on a transient gene expression assay, the function of two GFP genes was tested: mGFP-4 (Haseloff et al., 1997) and smRS-GFP (Davis and Vierstra, 1998). The intensity of smRS-GFP fluorescence was higher than that of mGFP-4 in Petunia and easier to distinguish from other unspecific fluorescent signals.
First stable transformation experiments in four commercially relevant Petunia cultivars involved the vector pMen65smRS-GFP containing nptII gene.
This vector resulted in clearly detectable GFP expressing callus and made it possible to visually select exclusively for GFP. Selection using GFP only versus GFP and kanamycin selection was compared regarding the transformation efficiencies.
The detection of integration the transgene via southern hybridization revealed single and multiple integrations of smRS-GFP in Petunia. Single copy plants showed intensive expression in all parts of the plants, while rising copy numbers led to only weak or partial expression of smRS-GFP. In this study, the possibility to select transgenic plants based on their GFP expression without applying antibiotic or herbicide resistance genes is shown.
Publication
Authors
V. Mußmann, M. Serek , T. Winkelmann
Keywords
Agrobacterium tumefaciens, GFP, Petunia hybrid, selection marker
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