Functional expression of the taste‐modifying protein, miraculin, in transgenic lettuce

Sun, Hyeon-Jin; Cui, Ming; Ma, Biao; Ezura, Hiroshi

doi:10.1016/j.febslet.2005.12.080

Cited by 115 publications

(109 citation statements)

References 31 publications

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“…In brief miraculin encoding 660-bp DNA fragment was inserted into the XbaI/SacI sites of the plant transformation vectors pBI121 (Sun et al 2006). Line 56B possesses a single copy of the miraculin gene driven by the CaMV 35S promoter.…”

Section: Plant Materials and Growth Conditionsmentioning

confidence: 99%

“…Nevertheless, commercial production of miraculin is limited because the natural source is a tropical plant that is difficult to cultivate and is not very productive. Efforts to produce miraculin in other organisms such as Escherichia coli (Kurihara 1992), yeast (Kurihara and Nirasawa 1997), tobacco (Kurihara and Nirasawa 1997), lettuce (Sun et al 2006), tomato (Sun et al 2007), and strawberry (Sugaya et al 2008) have been going on since 1990. Recently, a research group reattempted to produce miraculin in E. coli, but the recombinant protein exhibited lower taste-modifying activity than the native miraculin because the recombinant miraculin was not glycosylated (Matsuyama et al 2009).…”

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confidence: 99%

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Gene dosage and genetic background affect miraculin accumulation in transgenic tomato fruits

Kim

Hirai

Kato

et al. 2010

Plant Biotechnology

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Gene dosage and genetic background are factors that influence transgene expression in transgenic plants. In our previous studies, we produced transgenic tomato plants that accumulate miraculin, a taste-modifying protein, in a genetically stable manner. To elucidate the effects of gene dosage and genetic background on miraculin accumulation in transgenic tomato fruits, we generated hybrid tomato lines between the homozygous transgenic line 56B (background cultivar 'Moneymaker') and the pure cultivars 'Micro-Tom,' 'Moneymaker,' 'Ailsa Craig,' 'M82,' 'Rutgers' and 'Aichi-first' and analyzed them for miraculin mRNA expression and miraculin protein accumulation. The hybrid lines exhibited variation in their fruit structures. Among the hybrid lines heterozygous for the miraculin gene, miraculin accumulation in the fruits varied from 111.0 mg g Ϫ1 fresh weight (FW) to 159.4 mg g Ϫ1 FW. Furthermore, the homozygous line 56B showed higher miraculin accumulation and miraculin mRNA expression than the heterozygous line 56Bϫ'Moneymaker.' These results demonstrate the profound effects of gene dosage and genetic background on miraculin accumulation in transgenic tomato fruits.Key words: Gene dosage, genetic background, miraculin, taste-modifying protein.Plant Biotechnology 27, 333-338 (2010) Original PaperThis article can be found at http://www.jspcmb.jp/ allelic composition and genetic background affect transgene expression and inheritance in white clover. Another study in rice showed that the expression pattern of Xa3/Xa26, which is a rice disease resistance gene, was affected by the genetic background (Cao et al. 2007). The full effects of gene dosage and genetic background on transgene expression and translation remain to be elucidated.In this study, we generated hybrid lines between transgenic line 56B (background cultivar 'Moneymaker'), which is homozygous for the miraculin gene, and the pure line cultivars 'Micro-Tom,' 'Moneymaker,' 'Ailsa Craig,' 'M82,' 'Rutgers' and 'Aichi-first.' Characterization of the hybrids' miraculin mRNA expression and miraculin protein accumulation, revealed the effects of gene dosage and genetic background on miraculin accumulation in transgenic tomato fruits, and in addition we discuss the significance of combining molecular breeding with cross-breeding to produce transgenic tomatoes that accumulate miraculin. Materials and methods Plant material and growth conditionsThe transgenic tomato line 56B was produced in our previous work (Sun et al. 2007). In brief miraculin encoding 660-bp DNA fragment was inserted into the XbaI/SacI sites of the plant transformation vectors pBI121 (Sun et al. 2006). Line 56B possesses a single copy of the miraculin gene driven by the CaMV 35S promoter.In the present study, the miraculin-accumulating transgenic tomato line 56B (background cultivars 'Moneymaker') was crossed with six other cultivars: 'Micro-Tom (accession number TOMJPF00001), ' 'Moneymaker (accession number TOMJPF00002), ' 'Aichi-first (accession number TOMJPF00003), ' 'Ailsa Cra...

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Section: Plant Materials and Growth Conditionsmentioning

confidence: 99%

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confidence: 99%

Gene dosage and genetic background affect miraculin accumulation in transgenic tomato fruits

Kim

Hirai

Kato

et al. 2010

Plant Biotechnology

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“…Recently, however, when Aspergillus oryzae was used as a host for expressing miraculin, the recombinant protein did exhibit taste-modifying properties (Ito et al 2007). We have also successfully expressed recombinant miraculin in seedpropagated crop species, including lettuce (Sun et al 2006) and tomato (Sun et al 2007); however, in subsequent generations, stable miraculin expression was observed in tomato but silencing of the transgene occurred in lettuce, although the recombinant protein was correctly folded in both cases. This suggests that the plant species used for transformation and the mode of seedling propagation are important for producing miraculin-expressing transgenic plants.…”

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Transgenic strawberry expressing the taste-modifying protein miraculin

Sugaya

Yano

Sun

et al. 2008

Plant Biotechnology

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The gene encoding the taste-modifying protein miraculin was introduced under the control of the 35S or El2 promoter into strawberry (Fragaria x ananassa) by Agrobacterium-mediated transformation to produce transgenic plants. Although miraculin was detected in the leaves and fruits of the transgenic plants, the level of accumulation among the transgenic lines, which ranged from 0.5 to 2.0 mg g Ϫ1 fresh fruit, was not significantly different and was lower than that in miracle fruits (145 mg g Ϫ1 fresh fruit). High levels of miraculin accumulation were detected in the mature fruits. The transgenic lines were subsequently propagated via the runners for three vegetative generations, and miraculin was detected at equal levels in the leaves and fruits of the plants from each generation. In conclusion, although the level of accumulation was not high, miraculin was stably expressed and accumulated in the vegetative progeny of the transgenic strawberry plants.

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“…The content of recombinant VrPDF1 was determined by the ELISA method of Sun et al (2006) [23]. Soluble protein was extracted from T1 generation tobacco transformation seeds and diluted to a concentration of 200 µg ml -1 , and 100 µl VrPDF1 dilution was added into microplates three times.…”

Section: Analysis Of Recombinant Vrdef1 Protein By Western Blot and Ementioning

confidence: 99%

Expression analysis of recombinant Vigna radiata plant defensin 1 protein in transgenic tobacco plants

Thao¹,

Lan²,

Tường³

et al. 2017

J App Biol Biotech

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Bruchid resistance is regulated by defensin gene. Vigna radiata plant defensin 1 (VrPDF1) inhibits alpha-amylase activities in insect gut; therefore, insect will die of undigestion starch. VrPDF1 content in mungbean seeds is very low. Hence an increase in content of VrPDF1 in mungbean seeds leads to enhance alpha -amylase inhibition in larvae and bruchids, which is necessary for the study to improve bruchid resistance in mungbeans. This article presents the results of VrPDF1 gene expression in T1 generation transgenic tobacco seeds. It was confirmed that VrPDF1 gene was attached to genome of tobacco plants and translated to synthesize VrPDF1. Recombinant VrPDF1 protein was successfully expressed in seeds of four transgenic tobacco lines (T1 -7, T1-8, T1-10, T1-11), among which T1-10 line had the highest recombinant VrPDF1 content, reaching 8.57 g mg -1 total protein. The extract containing recombinant VrPDF1 from the transgenic tobacco lines effectively inhibited the activity of alpha-amylase from the intestine of larvae and weevils. However, the protein extract solution from T1 -10 line had the strongest inhibitory effect, so the activity of alpha-amylase was only 18.89% compared to controls. The analysis results of VrPDF1 gene expression in transgenic tobacco plants are fundamental to the transfer of pPhaso-dest-VrPDF1 vector into mungbean to improve bruchid resistance of mungbean and contribute to improving mungbean preservation.

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Functional expression of the taste‐modifying protein, miraculin, in transgenic lettuce

Cited by 115 publications

References 31 publications

Gene dosage and genetic background affect miraculin accumulation in transgenic tomato fruits

Gene dosage and genetic background affect miraculin accumulation in transgenic tomato fruits

Transgenic strawberry expressing the taste-modifying protein miraculin

Expression analysis of recombinant Vigna radiata plant defensin 1 protein in transgenic tobacco plants

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