Cloning an ipt gene from Agrobacterium tumefaciens: characterisation of cytokinins in derivative transgenic plant tissue

McKenzie, Marian J.; Jameson, Paula E.; Poulter, Russell T. M.

doi:10.1007/bf00024796

Cited by 15 publications

(11 citation statements)

References 43 publications

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“…For construction of the transformation vector, attR recombination sites (Gateway Technology, Invitrogen) were cloned into the vector pMJM (McKenzie et al, 1994), creating pHJMGW. A 4-kb-long cassette from pMJMGW containing the maize (Zea mays) ubiquinone promoter (ubi1) and the attR recombination sites was cloned into pCAMBIA2301.…”

Section: Agrobacterium Tumefaciens Strain and Culture Preparationmentioning

confidence: 99%

A Bifunctional Geranyl and Geranylgeranyl Diphosphate Synthase Is Involved in Terpene Oleoresin Formation inPicea abies

et al. 2009

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The conifer Picea abies (Norway spruce) defends itself against herbivores and pathogens with a terpenoid-based oleoresin composed chiefly of monoterpenes (C10) and diterpenes (C20). An important group of enzymes in oleoresin biosynthesis are the short-chain isoprenyl diphosphate synthases that produce geranyl diphosphate (C10), farnesyl diphosphate (C15), and geranylgeranyl diphosphate (C20) as precursors of different terpenoid classes. We isolated a gene from P. abies via a homology-based polymerase chain reaction approach that encodes a short-chain isoprenyl diphosphate synthase making an unusual mixture of two products, geranyl diphosphate (C10) and geranylgeranyl diphosphate (C20). This bifunctionality was confirmed by expression in both prokaryotic (Escherichia coli) and eukaryotic (P. abies embryogenic tissue) hosts. Thus, this isoprenyl diphosphate synthase, designated PaIDS1, could contribute to the biosynthesis of both major terpene types in P. abies oleoresin. In saplings, PaIDS1 transcript was restricted to wood and bark, and transcript level increased dramatically after methyl jasmonate treatment, which induces the formation of new (traumatic) resin ducts. Polyclonal antibodies localized the PaIDS1 protein to the epithelial cells surrounding the traumatic resin ducts. PaIDS1 has a close phylogenetic relationship to single-product conifer geranyl diphosphate and geranylgeranyl diphosphate synthases. Its catalytic properties and reaction mechanism resemble those of conifer geranylgeranyl diphosphate synthases, except that significant quantities of the intermediate geranyl diphosphate are released. Using site-directed mutagenesis and chimeras of PaIDS1 with single-product geranyl diphosphate and geranylgeranyl diphosphate synthases, specific amino acid residues were identified that alter the relative composition of geranyl to geranylgeranyl diphosphate.

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Section: Agrobacterium Tumefaciens Strain and Culture Preparationmentioning

confidence: 99%

A Bifunctional Geranyl and Geranylgeranyl Diphosphate Synthase Is Involved in Terpene Oleoresin Formation inPicea abies

et al. 2009

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“…Two primers were designed that flanked the coding and termination regions of a previously isolated ipt gene sequence (McKenzie et al, 1994). PCR was performed using a Perkin Elmer-Cetus kit according to the manufacturer's instructions.…”

Section: Pcrmentioning

confidence: 99%

“…This gene encodes the enzyme isopentenyl transferase, which catalyzes the rate-limiting step of the cytokinin biosynthetic pathway (Akiyoshi et al, 1984;Barry et al, 1984), in which ⌬ 2 -isopentenyl PPi is condensed with AMP to form isopentenyl AMP. The introduction of the ipt gene into the plant genome results in elevated levels of cytokinin in the transformed tissue (Smigocki and Owens, 1988;Beinsberger et al, 1991;Yusibov et al, 1991;McKenzie et al, 1994), together with associated morphological changes.…”

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confidence: 99%

“…The altered morphology observed in plant tissues transformed with the native ipt gene has been clearly associated with a marked increase in cytokinin content (Budar et al, 1986;Smigocki and Owens, 1988;Beinsberger et al, 1991;Yusibov et al, 1991;McKenzie et al, 1994). Additional work has been carried out using the CaMV 35S promoter to drive ipt gene expression (Smigocki andOwens, 1988, 1989).…”

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confidence: 99%

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Controlled Cytokinin Production in Transgenic Tobacco Using a Copper-Inducible Promoter

et al. 1998

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The cytokinin group of plant hormones regulates aspects of plant growth and development, including the release of lateral buds from apical dominance and the delay of senescence. In this work the native promoter of a cytokinin synthase gene (ipt) was removed and replaced with a Cu-controllable promoter. Tobacco (Nicotiana tabacum L. cv tabacum) transformed with this Cu-inducible ipt gene (Cu-ipt) was morphologically identical to controls under noninductive conditions in almost all lines produced. However, three lines grew in an altered state, which is indicative of cytokinin overproduction and was confirmed by a full cytokinin analysis of one of these lines. The in vitro treatment of morphologically normal Cu-ipt transformants with Cu 2؉ resulted in delayed leaf senescence and an increase in cytokinin concentration in the one line analyzed. In vivo, inductive conditions resulted in a significant release of lateral buds from apical dominance. The morphological changes seen during these experiments may reflect the spatial aspect of control exerted by this gene expression system, namely expression from the root tissue only. These results confirmed that endogenous cytokinin concentrations in tobacco transformants can be temporally and spatially controlled by the induction of ipt gene expression through the Cu-controllable gene-expression system.

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“…In those transgenic plants, the ipt gene was driven by various promoters, for example constitutive, native, and inducible promoters (Smigocki 1991;Yusibov et al 1991;Rupp et al 1999;van der Graaff et al 2001;Geng et al 2001Geng et al , 2002. Transgenic plants or tissues with ipt expression under the control of a constitutive promoter have been clearly associated with a remarkable increase in cytokinin content; this increase, in turn, resulted in dramatic changes in morphological characteristics, such as non-elongated shoots, bushy phenotypes with a severe lack of apical dominance, small round or serrated leaves, undeveloped roots, delayed leaf senescence, and sterility (Smigocki 1991;Mckonzie et al 1994;Geng et al 2001;van der Graaff et al 2001;Guo et al 2005). Although a number of studies have found that cytokinins have a clear role in plant growth and cell development, it remains to be elucidated how the altered in-vivo hormonal concentrations can vigorously trigger plant development.…”

Section: Introductionmentioning

confidence: 99%

Isopentenyl transferase gene (ipt) downstream transcriptionally fused with gene expression improves the growth of transgenic plants

Guo

Duan

et al. 2009

Transgenic Res

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This research reports a promising approach to increase a plant's physiological cytokinin content. This approach also enables the increase to play a role in plant growth and development by introducing the ipt gene to downstream transcriptionally fuse with other genes under the control of a CaMV35S promoter, in which the ipt gene is far from the 35S promoter. According to Kozak's ribosome screening model, expression of the ipt gene is reduced by the terminal codon of the first gene and the internal untranslated nucleotides between the fused genes. In the transgenic plants pVKH35S-GUS-ipt, pVKH35S-AOC-ipt, and pVKH35S-AtGolS2-ipt, cytokinins were increased only two to threefold, and the plants grew more vigorously than the pVKH35S-AOC or pVKH35S-AtGolS2 transgenic plants lacking the ipt gene. The vigorous growth was reflected in rapid plant growth, a longer flowering period, a greater number of flowers, more seed product, and increased chlorophyll synthesis. The AOC and AtGolS2 genes play a role in a plant's tolerance of salt or cold, respectively. When the ipt gene transcriptionally fuses with AOC or AtGolS2 in the frame of AOC-ipt and AtGolS2-ipt, slight cytokinin increases were obtained in their transgenic plants; furthermore, those increases played a positive role in improvements of plant growth. Notably, an increased cytokinin volume at the physiological level, in concert with AtGolS2 expression, enhances a plant's tolerance to cold.

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Cloning an ipt gene from Agrobacterium tumefaciens: characterisation of cytokinins in derivative transgenic plant tissue

Cited by 15 publications

References 43 publications

A Bifunctional Geranyl and Geranylgeranyl Diphosphate Synthase Is Involved in Terpene Oleoresin Formation inPicea abies

A Bifunctional Geranyl and Geranylgeranyl Diphosphate Synthase Is Involved in Terpene Oleoresin Formation inPicea abies

Controlled Cytokinin Production in Transgenic Tobacco Using a Copper-Inducible Promoter

Isopentenyl transferase gene (ipt) downstream transcriptionally fused with gene expression improves the growth of transgenic plants

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