Green fluorescent protein as a marker for expression of a second gene in transgenic plants

Harper, Brian K.; Mabon, Stephen A.; Leffel, S. M.; Halfhill, Matthew D.; Richards, H. A.; Moyer, K. A.; Stewart, C. Neal

doi:10.1038/15114

Cited by 109 publications

(69 citation statements)

References 26 publications

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“…The advent of fluorescent proteins, such as green fluorescent protein (GFP) from the jellyfish Aequorea victoria (Chalfie et al ., 1994) and its variants, has endowed researchers with powerful tools to monitor gene expression (Harper et al ., 1999;Sunilkumar et al , 2002) and gene silencing (Ruiz et al ., 1998), and to define spatial (Johnson et al ., 2005), developmental (Marton et al ., 2005) and quantitative (Tang & Newton, 2004) properties of promoters. The lack of requirement for exogenous substrate, cofactors or histochemical fixation makes GFP a particularly valuable reporter for plant cells, which are not as permeable as animal cells (Hanson & Köhler, 2001).…”

Section: Introductionmentioning

confidence: 99%

The dark side of green fluorescent protein

et al. 2005

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Summary• Here, severe interference of chlorophyll with green fluorescent protein (GFP) fluorescence is described for medicago ( Medicago truncatula ), rice ( Oryza sativa ) and arabidopsis ( Arabidopsis thaliana ). This interference disrupts the proportional relationship between GFP content and fluorescence that is intrinsic to its use as a quantitative reporter.• The involvement of chlorophyll in the loss of GFP fluorescence with leaf age was shown in vivo , by the removal of chlorophyll through etiolation or by ethanol extraction, and in vitro , by titration of a GFP solution with chlorophyll solutions of various concentrations.• A substantial decrease in fluorescence in early development of medicago and rice leaves correlated with chlorophyll accumulation. In all three species tested, removal of chlorophyll yielded up to a 10-fold increase in fluorescence. Loss of GFP fluorescence in vitro was 4-fold greater for chlorophyll b than for chlorophyll a.• Differences exist between plant species for the discrepancy between apparent GFP fluorescence and its actual level in green tissues. Substantial errors in estimating promoter activity from GFP fluorescence can occur if pigment interference is not considered.

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Section: Introductionmentioning

confidence: 99%

The dark side of green fluorescent protein

et al. 2005

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“…However, overexpression of GFP was reported to be toxic to plant growth and development (Haseloff et al, 1997), which we did not find in our present study. This attribute may depend on the particular GFP (Harper et al, 1999) and host plant used.…”

Section: Discussionmentioning

confidence: 99%

“…Researchers have been fascinated by the use of GFP genes as markers in many hosts (Oparka et al, 1997;Harper et al, 1999;Harper and Stewart, 2000;Hudson et al, 2001). Various transgenic GFP genes have been developed (Halfhill et al, 2003), and one of them actually was used as a marker gene to detect GM plants in field experiments .…”

Section: Discussionmentioning

confidence: 99%

Development of Visible Markers for Transgenic Plants and their Availability for Environmental Risk Assessment

Tamaoki

Imai²,

Takahashi³

et al. 2006

Zeitschrift Für Naturforschung C

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Monitoring of transgenic plants in the field is important, but risk assessment has entailed laborious use of invisible marker genes. Here, we assessed three easily visible marker transgenes Ð green fluorescent protein (GFP), R, and Nicotiana tabacum homeobox (NTH) 15 genes Ð for their potential use as marker genes for monitoring genetically modified plants. Transgenic Arabidopsis thaliana plants for each of these genes were visibly distinguished from wild-type plants. We determined the germination rate, 3-week fresh weight, time to first flowering, and seed weight of the transgenic plants to evaluate whether the expression of these marker genes affected the growth of the host. Introduction of GFP gene had no effect on the evaluated parameters, and we then used the GFP gene as a marker to assess the outcrossing frequency between transgenic and two Arabidopsis species. Our results showed that the hybridization frequency between transgenic plants and Arabidopsis thaliana was 0.24%, and between transformants and Arabidopsis lyrata it was 2.6% under experimental condition. Out-crossing frequency was decreased by extending the distance between two kinds of plants. Thus, the GFP gene is a useful marker for assessing the whereabouts of transgenes/transformants in the field. We also demonstrated that the GFP gene is possibly applicable as a selection marker in the process of generation of transgenic plants.

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“…GFP can also serve as a gene monitoring system that can be used to study the introgression of transgenes into wild relatives through hybridization, backcrossing, and transgene escape [156,157].…”

Section: Release Of Recombinant Proteins Other Than Cry Proteins In Rmentioning

confidence: 99%

Fate and Effects in Soil of Cry Proteins from Bacillus thuringiensis: Influence of Physicochemical and Biological Characteristics of Soil

Saxena¹,

Pushalkar

Stotzky³

2013

TOTNJ

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Bacillus thuringiensis (Bt) is a useful alternative or supplement to synthetic chemical pesticides in agriculture, forest management, and control of mosquitoes and some other biting insects. When modified Bt cry genes are inserted into a plant species (e.g., corn, cotton, potato, canola, rice), the plant expresses active larvicidal proteins in its tissues. The toxins continue to be synthesized during growth of the plants, making the plant toxic to various insect pests throughout their life or as biomass incorporated into soil. If production exceeds consumption, inactivation, and degradation, the toxins could accumulate to concentrations that may enhance the control of target pests or constitute a hazard to nontarget organisms, such as the soil microbiota, beneficial insects (e.g., pollinators, predators and parasites of insect pests), and other animal classes. The accumulation and persistence of the toxins could also result in the selection and enrichment of toxin-resistant target insects. Persistence is enhanced when the toxins are bound on surface-active particles in the environment (e.g., clays and humic substances) and, thereby, rendered more resistant to biodegradation while retaining toxic activity. Moreover, major problem we face today is of "Molecular pharming" that utilizes transgenic plants and animals for production of pharmaceuticals and chemicals for their use in human beings and industries respectively. Their release to the environment, especially to soil and potentially to waters of the pharmaceutical and industrial products of transgenic plant and animal "pharms" could pose a hazard to the environment. In contrast to the products of most transgenic plants currently available commercially (e.g., the insecticidal proteins from subspecies of Bt) that primarily target insects and other pests. These "pharms" are being genetically engineered to express products for use primarily in human beings. Consequently, these products constitute a class of compounds that is seldom found in natural habitats and that primarily target "higher level" eukaryotes. Hence, they are xenobiotics with respect to the environment, and their persistence in and effects on the environment have not been adequately studied and sober risk assessments on a case-bycase basis must be made before major releases of such transgenic organisms.

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Green fluorescent protein as a marker for expression of a second gene in transgenic plants

Cited by 109 publications

References 26 publications

The dark side of green fluorescent protein

The dark side of green fluorescent protein

Development of Visible Markers for Transgenic Plants and their Availability for Environmental Risk Assessment

Fate and Effects in Soil of Cry Proteins from Bacillus thuringiensis: Influence of Physicochemical and Biological Characteristics of Soil

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