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Coppoolse, Eric R.; Vroomen, Marianne J. de; Roelofs, Dick; Smit, Jan Maerten; Gennip, Femke van; Hersmus, Bart J. M.; Nijkamp, H. J. J.; Haaren, M. J. J. Van

doi:10.1023/a:1021174726070

Cited by 86 publications

(19 citation statements)

References 42 publications

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“…No corresponding bands were obtained in the case of untransformed control line. The transgenic lines grew normally and did not show any deleterious effect due to the presence of the cre driven by a CaMV35S promoter gene as reported earlier by Coppoolse et al (2003). When transferred to the soil, they flowered and set seed.…”

Section: Construct Designing Transformation and Selection Of A Thalsupporting

confidence: 77%

Generation of marker free salt tolerant transgenic plants of Arabidopsis thaliana using the gly I gene and cre gene under inducible promoters

Roy

Saxena

Bhomkar

et al. 2008

Plant Cell Tiss Organ Cult

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Despite the advances in transgenesis, transformation technologies still rely on the introduction of a selectable marker gene to identify cells and tissues that have integrated the gene of interest in their genome. The continuous presence of the marker genes in the transgenics is often controversial as it can potentially have multiple undesirable impacts. The present study employed the self-excising Cre-loxP system to generate marker-free Arabidopsis thaliana expressing the agronomically important glyoxalase I (glyI) gene from Brassica juncea to confer salt stress tolerance. A binary vector was constructed wherein the salt-inducible rd29A promoter was used to drive the expression of the glyI gene and the transformants of A. thaliana were recovered using kanamycin resistance as the selectable marker. The neomycin phosphotransferase II (nptII) gene was flanked by the loxP sites followed by the introduction of a heatinducible Cre-recombinase in between the loxP sites. The kanamycin-resistant transgenic lines of A. thaliana using this vector showed an ability to withstand stress imposed by 150 mM NaCl. The exposure of the T2 transgenic lines to a mild heat shock (37°C) resulted in the recovery of salt-tolerant, kanamycinsensitive T3 progeny. Molecular analyses of the T3 transgenic lines following the heat shock treatment confirmed the excision of the nptII gene and the completion of their life cycle in the presence of 150 mM NaCl-induced stress.

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Section: Construct Designing Transformation and Selection Of A Thalsupporting

confidence: 77%

Generation of marker free salt tolerant transgenic plants of Arabidopsis thaliana using the gly I gene and cre gene under inducible promoters

Roy

Saxena

Bhomkar

et al. 2008

Plant Cell Tiss Organ Cult

View full text Add to dashboard Cite

show abstract

“…However, retransformation is lengthy and cross-fertilization-dependent strategies are applicable only for sexually propagated species with relatively short reproduction cycles. Additionally, constitutive overexpression of recombinase proteins may cause aberrant developmental phenotypes in plants (Sonti et al 1995;Que et al 1998;Coppoolse et al 2003). Therefore, alternative approaches based on transient gene expression have been developed.…”

Section: Introductionmentioning

confidence: 99%

Site-specific excisional recombination strategies for elimination of undesirable transgenes from crop plants

Gidoni

Srivastava

Carmi

2008

In Vitro Cell.Dev.Biol.-Plant

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A major limitation of crop biotechnology and breeding is the lack of efficient molecular technologies for precise engineering of target genomic loci. While transformation procedures have become routine for a growing number of plant species, the random introduction of complex transgenenic DNA into the plant genome by current methods generates unpredictable effects on both transgene and homologous native gene expression. The risk of transgene transfer into related plant species and consumers is another concern associated with the conventional transformation technologies. Various approaches to avoid or eliminate undesirable transgenes, most notably selectable marker genes used in plant transformation, have recently been developed. These approaches include cotransformation with two independent T-DNAs or plasmid DNAs followed by their subsequent segregation, transposon-mediated DNA elimination, and most recently, attempts to replace bacterial T-DNA borders and selectable marker genes with functional equivalents of plant origin. The use of site-specific recombination to remove undesired DNA from the plant genome and concomitantly, via excision-mediated DNA rearrangement, switch-activate by choice transgenes of agronomical, food or feed quality traits provides a versatile "transgene maintenance and control" strategy that can significantly contribute to the transfer of transgenic laboratory developments into farming practice. This review focuses on recent reports demonstrating the elimination of undesirable transgenes (essentially selectable marker and recombinase genes) from the plant genome and concomitant activation of a silent transgene (e.g., a reporter gene) mediated by different site-specific recombinases driven by constitutive or chemically, environmentally or developmentally regulated promoters. These reports indicate major progress in excision strategies which extends application of the technology from annual, sexually propagated plants towards perennial, woody and vegetatively propagated plants. Current trends and future prospects for optimization of excision-activation machinery and its practical implementation for the generation of transgenic plants and plant products free of undesired genes are discussed.

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“…Two types of effects have been described: phenotypic abnormalities and DNA rearrangements in chloroplasts. In petunia, tomato, tobacco and Arabidopsis aberrant phenotypes such as leaf chlorosis, growth retardation and reduced fertility were associated with high levels of cre expression (Coppoolse et al, 2003). These phenotypic abnormalities were not connected with chromosomal rearrangements: they always co-segregated with the cre transgene.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, additional time is required to perform a second round of transformation or cross pollination. A further strong argument against constitutive cre expression is the possible occurrence of genetic and phenotypic changes caused by the Cre recombinase, which were observed in plastid and nuclear genomes, respectively (Hajdukiewicz et al, 2001;Coppoolse et al, 2003).…”

Section: Cre Expression Strategies: Efficiency and Limitationsmentioning

confidence: 99%