Bundoora, Australia
Bundoora, Australia

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Brugliera F.,Florigene Pty Ltd. | Brugliera F.,La Trobe University | Tao G.-Q.,Florigene Pty Ltd. | Tao G.-Q.,La Trobe University | And 14 more authors.
Plant and Cell Physiology | Year: 2013

Chrysanthemums (Chrysanthemum×morifolium Ramat.) are an important cut-flower and potted plant crop in the horticultural industry world wide. Chrysanthemums express the flavonoid 3′-hydroxylase (F3′H) gene and thus accumulate anthocyanins derived from cyanidin in their inflorescences which appear pink/red. Delphinidin-based anthocyanins are lacking due to the deficiency of a flavonoid 3′, 5′-hydroxylase (F3′5′H), and so violet/blue chrysanthemum flower colors are not found. In this study, together with optimization of transgene expression and selection of the host cultivars and gene source, F3′5′H genes have been successfully utilized to produce transgenic bluish chrysanthemums that accumulate delphinidin-based anthocyanins. HPLC analysis and feeding experiments with a delphinidin precursor identified 16 cultivars of chrysanthemums out of 75 that were predicted to turn bluish upon delphinidin accumulation. A selection of eight cultivars were successfully transformed with F3′5′H genes under the control of different promoters. A pansy F3′5′H gene under the control of a chalcone synthase promoter fragment from rose resulted in the effective diversion of the anthocyanin pathway to produce delphinidin in transgenic chrysanthemum flower petals. The resultant petal color was bluish, with 40% of total anthocyanidins attributed to delphinidin. Increased delphinidin levels (up to 80%) were further achieved by hairpin RNA interference-mediated silencing of the endogenous F3′H gene. The resulting petal colors were novel bluish hues, not possible by hybridization breeding. This is the first report of the production of anthocyanins derived from delphinidin in chrysanthemum petals leading to novel flower color. © 2013 The Author.


Nakamura N.,Suntory Holdings Ltd. | Tems U.,Florigene Pty Ltd | Fukuchi-Mizutani M.,Suntory Holdings Ltd. | Chandler S.,Florigene Pty Ltd | And 4 more authors.
Plant Biotechnology | Year: 2011

An important part of the assessment of the potential environmental impact from the introduction of a genetically modified (GM) plant is an evaluation of the potential for gene flow from the GM plant to related wild species. This information is needed as part of the risk-assessment process, in the context of whether gene flow to wild species is possible. One method for evaluating gene flow is to use molecular techniques to identify genes in wild species populations that may have originated from a cultivated species. An advantage of this method is that a phenotypic marker or trait is not required to measure gene flow. In the present study we analyzed the seedlings of seeds from three wild native Rosa species (R. multiflora Thunb., R. luciae Rochebr. et Franch. ex Crép. and R. rugosa Thunb.) selected from several locations across Japan where the wild rose was growing in close proximity to cultivated rose plants (Rosxhybrida). To determine whether gene flow from cultivated rose had occurred, young leaves of 1,296 seedlings from the wild Rosa plants were analyzed by PCR for the presence of the KSN locus. This locus originated from a sport of R. chinensis Jacq. var. spontanea (Rehd. Et Wils.) Yu et Ku and is involved in the recurrent flowering phenotype observed for cultivated rose hybrids, but is absent in Japanese species roses. The KSN locus was absent in all seedlings sampled, indicating no gene flow to wild Rosa species from the cultivated rose had occurred, and providing evidence that the probability of gene flow from cultivated to wild Rosa species in Japan is low or non-existent.


Nakamura N.,Suntory Holdings Ltd | Fukuchi-Mizutani M.,Suntory Holdings Ltd | Katsumoto Y.,Suntory Holdings Ltd | Togami J.,Suntory Holdings Ltd | And 12 more authors.
Plant Biotechnology | Year: 2011

The release of genetically modified plants into the environment can only occur after permission is obtained from the relevant regulatory authorities. This permission will only be obtained after extensive risk assessment shows comparable risk of impact to the environment and biodiversity as compared to non-transgenic host plants. Two transgenic rose (Rosa×hybrida) lines, whose flowers were modified to a bluer colour as a result of accumulation of delphinidin-based anthocyanins, have been trialed in greenhouses and the field in both Japan and Australia. Flower colour modification was due to expression of genes of a viola flavonoid 3',5'-hydroxylase and a torenia anthocyanin 5-acyltransferase. In all trials it was shown that the performance of the two transgenic lines, as measured by their growth characters, was comparable to the host untransformed variety. Biological assay showed that the transgenic lines did not produce allelopathic compounds. In Japan, seeds from wild rose species that had grown in close proximity to the transgenic roses did not carry either a Rosa×hybrida specific marker gene or the transgenes. In hybridization experiments using transgenic rose pollen and wild rose female parents, the transgenes were not detected in the seed obtained, though there was a low frequency of seed set. The transgene was also not transmitted when Rosa×hybrida cultivars were used as females. In in situ hybridization analysis transgene transcripts were only detected in the epidermal cells in the petals of the transgenic roses. In combination, the breeding and in situ analysis results show that the transgenic roses contain the transgene only in the L1 layer cells and not in the L2 layer cells that generate reproductive cells. General release permissions have been granted for both transgenic lines in Japan and one is now commercially produced.


Tanaka Y.,Suntory Holdings Ltd. | Brugliera F.,Florigene Pty Ltd. | Kalc G.,Florigene Pty Ltd. | Senior M.,Florigene Pty Ltd. | And 4 more authors.
Bioscience, Biotechnology and Biochemistry | Year: 2010

The status quo of flavonoid biosynthesis as it relates to flower color is reviewed together with a success in modifying flower color by genetic engineering. Flavo-noids and their colored class compounds, anthocyanins, are major contributors to flower color. Many plant species synthesize limited kinds of flavonoids, and thus exhibit a limited range of flower color. Since genes regulating flavonoid biosynthesis are available, it is possible to alter flower color by overexpressing heter-ologous genes and/or down regulating endogenous genes. Transgenic carnations and a transgenic rose that accumulate delphinidin as a result of expressing a flavonoid 3',5'-hydroxylase gene and have novel blue hued flowers have been commercialized. Transgenic Nierembergia accumulating pelargonidin, with novel pink flowers, has also been developed. Although it is possible to generate white, yellow, and pink-flowered torenia plants from blue cultivars by genetic engineering, field trial observations indicate difficulty in obtaining stable phenotypes.


Chandler S.F.,Florigene Pty. Ltd. | Brugliera F.,Florigene Pty. Ltd.
Biotechnology Letters | Year: 2011

Micro-propagation, embryo rescue, mutagenesis via chemical or irradiation means and in vitro inter-specific hybridisation methods have been used by breeders in the floriculture industry for many years. In the past 20 years these enabling technologies have been supplemented by genetic modification methods. Though many genes of potential utility to the floricultural industry have been identified, and much has been learnt of the genetic factors and molecular mechanisms underlying phenotypes of great importance to the industry, there are only flower colour modified varieties of carnation and rose in the marketplace. To a large extent this is due to unique financial barriers to market entry for genetically modified varieties of flower crops, including use of technology fees and costs of regulatory approval. © 2010 Springer Science+Business Media B.V.


Tanaka Y.,Suntory Business Expert Ltd | Brugliera F.,Florigene Pty Ltd
Philosophical Transactions of the Royal Society B: Biological Sciences | Year: 2013

Cytochromes P450 play important roles in biosynthesis of flavonoids and their coloured class of compounds, anthocyanins, both of which are major floral pigments. The number of hydroxyl groups on the B-ring of anthocyanidins (the chromophores and precursors of anthocyanins) impact the anthocyanin colour, the more the bluer. The hydroxylation pattern is determined by two cytochromes P450, flavonoid 30-hydroxylase (F30H) and flavonoid 30,50-hydroxylase (F3050H) and thus they play a crucial role in the determination of flower colour. F30H and F3050H mostly belong to CYP75B and CYP75A, respectively, except for the F3050Hs in Compositae that were derived from gene duplication of CYP75B and neofunctionalization. Roses and carnations lack blue/violet flower colours owing to the deficiency of F3050H and therefore lack the B-ring-trihydroxylated anthocyanins based upon delphinidin. Successful redirection of the anthocyanin biosynthesis pathway to delphinidin was achieved by expressing F3050H coding regions resulting in carnations and roses with novel blue hues that have been commercialized. Suppression of F3050H and F30H in delphinidinproducing plants reduced the number of hydroxyl groups on the anthocyanidin B-ring resulting in the production of monohydroxylated anthocyanins based on pelargonidin with a shift in flower colour to orange/red. Pelargonidin biosynthesis is enhanced by additional expression of a dihydroflavonol 4-reductase that can use the monohydroxylated dihydrokaempferol (the pelargonidin precursor). Flavone synthase II (FNSII)-catalysing flavone biosynthesis from flavanones is also a P450 (CYP93B) and contributes to flower colour, because flavones act as co-pigments to anthocyanins and can cause blueing and darkening of colour. However, transgenic plants expression of a FNSII gene yielded paler flowers owing to a reduction of anthocyanins because flavanones are precursors of anthocyanins and flavones. © 2013 The Author(s) Published by the Royal Society. All rights reserved.


Gion K.,Suntory Business Expert Ltd. | Suzuri R.,Suntory Business Expert Ltd. | Ishiguro K.,Suntory Business Expert Ltd. | Katsumoto Y.,Suntory Business Expert Ltd. | And 5 more authors.
Acta Horticulturae | Year: 2012

We have been using genetic engineering to develop and commercialize new floricultural cultivars focusing on novel flower colour. Rose, carnation and chrysanthemum do not have violet or blue flowers due to their lack of ability to synthesize the delphinidin-based anthocyanins which most violet/blue flowers contain. Flavonoid 3', 5'-hydroxylase (F3'5'H) is the key enzyme on the biosynthesis pathway leading to delphinidin. Transgenic rose, carnation and chrysanthemum expressing a heterologous F3'5'H gene produced flowers with a novel violet/blue flower colour for these species, not possible by hybridization breeding. Transgenic carnations are sold in USA, EU and Japan and a transgenic rose in Japan. Rose, carnation and chrysanthemum expressing an Arabidopsis FT (a gene that promotes flowering) exhibited very early flowering phenotypes. Expressing dominant chimeric repressors of various transcriptional factors in transgenic rose resulted in various novel flower morphologies. These technologies will be useful for development of novel floricultural crops in future.


PubMed | Florigene Pty. Ltd.
Type: Journal Article | Journal: Biotechnology letters | Year: 2011

Micro-propagation, embryo rescue, mutagenesis via chemical or irradiation means and in vitro inter-specific hybridisation methods have been used by breeders in the floriculture industry for many years. In the past 20 years these enabling technologies have been supplemented by genetic modification methods. Though many genes of potential utility to the floricultural industry have been identified, and much has been learnt of the genetic factors and molecular mechanisms underlying phenotypes of great importance to the industry, there are only flower colour modified varieties of carnation and rose in the marketplace. To a large extent this is due to unique financial barriers to market entry for genetically modified varieties of flower crops, including use of technology fees and costs of regulatory approval.


PubMed | Florigene Pty Ltd.
Type: Journal Article | Journal: Plant & cell physiology | Year: 2013

Chrysanthemums (Chrysanthemummorifolium Ramat.) are an important cut-flower and potted plant crop in the horticultural industry world wide. Chrysanthemums express the flavonoid 3-hydroxylase (F3H) gene and thus accumulate anthocyanins derived from cyanidin in their inflorescences which appear pink/red. Delphinidin-based anthocyanins are lacking due to the deficiency of a flavonoid 3, 5-hydroxylase (F35H), and so violet/blue chrysanthemum flower colors are not found. In this study, together with optimization of transgene expression and selection of the host cultivars and gene source, F35H genes have been successfully utilized to produce transgenic bluish chrysanthemums that accumulate delphinidin-based anthocyanins. HPLC analysis and feeding experiments with a delphinidin precursor identified 16 cultivars of chrysanthemums out of 75 that were predicted to turn bluish upon delphinidin accumulation. A selection of eight cultivars were successfully transformed with F35H genes under the control of different promoters. A pansy F35H gene under the control of a chalcone synthase promoter fragment from rose resulted in the effective diversion of the anthocyanin pathway to produce delphinidin in transgenic chrysanthemum flower petals. The resultant petal color was bluish, with 40% of total anthocyanidins attributed to delphinidin. Increased delphinidin levels (up to 80%) were further achieved by hairpin RNA interference-mediated silencing of the endogenous F3H gene. The resulting petal colors were novel bluish hues, not possible by hybridization breeding. This is the first report of the production of anthocyanins derived from delphinidin in chrysanthemum petals leading to novel flower color.

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