Pub Date : 2018-09-25DOI: 10.5511/PLANTBIOTECHNOLOGY.18.0515A
Shunji Nakamura, T. Oyama
The circadian system of plants is based on the cell-autonomously oscillating circadian clock. In the plant body, these cellular clocks are associated with each other, but their basic and intrinsic properties are still largely unknown. Here we report a method that enables long-term monitoring of bioluminescence circadian rhythms of a protoplast culture in a complete synthetic medium. From the leaves of Arabidopsis transgenic plants carrying the luciferase gene under a clock-gene promoter, mesophyll protoplasts were isolated and their bioluminescence was automatically measured every 20 min for more than one week. Decreasing luminescence intensities were observed in protoplasts when they were cultured in a Murashige and Skoog-based medium and also in W5 solution. This decrease was dramatically improved by adding the phytohormones auxin and cytokinin to the MS-based medium; robust circadian rhythms were successfully monitored. Interestingly, the period lengths of bioluminescence circadian rhythms of protoplasts under constant conditions were larger than those of detached leaves, suggesting that the period lengths of mesophyll cells in leaves were modulated from their intrinsic properties by the influence of other tissues/cells. The entrainability of protoplasts to light/dark signals was clearly demonstrated by using this monitoring system. By analyzing the circadian behavior of isolated protoplasts, the basic circadian system of plant cells may be better understood.
{"title":"Long-term monitoring of bioluminescence circadian rhythms of cells in a transgenic Arabidopsis mesophyll protoplast culture.","authors":"Shunji Nakamura, T. Oyama","doi":"10.5511/PLANTBIOTECHNOLOGY.18.0515A","DOIUrl":"https://doi.org/10.5511/PLANTBIOTECHNOLOGY.18.0515A","url":null,"abstract":"The circadian system of plants is based on the cell-autonomously oscillating circadian clock. In the plant body, these cellular clocks are associated with each other, but their basic and intrinsic properties are still largely unknown. Here we report a method that enables long-term monitoring of bioluminescence circadian rhythms of a protoplast culture in a complete synthetic medium. From the leaves of Arabidopsis transgenic plants carrying the luciferase gene under a clock-gene promoter, mesophyll protoplasts were isolated and their bioluminescence was automatically measured every 20 min for more than one week. Decreasing luminescence intensities were observed in protoplasts when they were cultured in a Murashige and Skoog-based medium and also in W5 solution. This decrease was dramatically improved by adding the phytohormones auxin and cytokinin to the MS-based medium; robust circadian rhythms were successfully monitored. Interestingly, the period lengths of bioluminescence circadian rhythms of protoplasts under constant conditions were larger than those of detached leaves, suggesting that the period lengths of mesophyll cells in leaves were modulated from their intrinsic properties by the influence of other tissues/cells. The entrainability of protoplasts to light/dark signals was clearly demonstrated by using this monitoring system. By analyzing the circadian behavior of isolated protoplasts, the basic circadian system of plant cells may be better understood.","PeriodicalId":20411,"journal":{"name":"Plant Biotechnology","volume":"35 3 1","pages":"291-295"},"PeriodicalIF":1.6,"publicationDate":"2018-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.5511/PLANTBIOTECHNOLOGY.18.0515A","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49035433","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-09-25DOI: 10.5511/PLANTBIOTECHNOLOGY.18.0510A
M. Wada, H. Oshino, N. Tanaka, N. Mimida, Y. Moriya-Tanaka, C. Honda, T. Hanada, H. Iwanami, S. Komori
Apple MdMADS13 has a transcription factor with MADS domain. Moreover, it is expressed specifically at petals and carpels. The product forms a dimer with MdPISTILLATA (MdPI) protein as a class B gene for floral organ formation. Reportedly, in parthenocarpic cultivars of apple (Spencer Seedless, Wellington Bloomless, Wickson and Noblow) the MdPI function is lost by genome insertion of retrotransposon, which cultivars show a homeotic mutation of floral organs, petals to sepals and stamens to carpels. Apple fruit is pome from receptacle tissue, and MdSEPALLATA (MdMADS8/9) and AGAMOUS homologues MdMADS15/22 involved in the fruit development, the transgenic apple suppressed these gene showed poor fruit development and abnormal flower formation. This article describes that the MdMADS13 retained expression after blossom and small fruits of parthenocarpic cultivars. Yeast two-hybrid experiment showed specific binding between MdPI and MdMADS13 proteins. Furthermore, transgenic Arabidopsis with 35S::MdMADS13 have malformed stamens and carpels. These results suggest strongly that MdMADS13 is related to flower organ formation as a class B gene with MdPI.
{"title":"Expression and functional analysis of apple MdMADS13 on flower and fruit formation.","authors":"M. Wada, H. Oshino, N. Tanaka, N. Mimida, Y. Moriya-Tanaka, C. Honda, T. Hanada, H. Iwanami, S. Komori","doi":"10.5511/PLANTBIOTECHNOLOGY.18.0510A","DOIUrl":"https://doi.org/10.5511/PLANTBIOTECHNOLOGY.18.0510A","url":null,"abstract":"Apple MdMADS13 has a transcription factor with MADS domain. Moreover, it is expressed specifically at petals and carpels. The product forms a dimer with MdPISTILLATA (MdPI) protein as a class B gene for floral organ formation. Reportedly, in parthenocarpic cultivars of apple (Spencer Seedless, Wellington Bloomless, Wickson and Noblow) the MdPI function is lost by genome insertion of retrotransposon, which cultivars show a homeotic mutation of floral organs, petals to sepals and stamens to carpels. Apple fruit is pome from receptacle tissue, and MdSEPALLATA (MdMADS8/9) and AGAMOUS homologues MdMADS15/22 involved in the fruit development, the transgenic apple suppressed these gene showed poor fruit development and abnormal flower formation. This article describes that the MdMADS13 retained expression after blossom and small fruits of parthenocarpic cultivars. Yeast two-hybrid experiment showed specific binding between MdPI and MdMADS13 proteins. Furthermore, transgenic Arabidopsis with 35S::MdMADS13 have malformed stamens and carpels. These results suggest strongly that MdMADS13 is related to flower organ formation as a class B gene with MdPI.","PeriodicalId":20411,"journal":{"name":"Plant Biotechnology","volume":"35 3 1","pages":"207-213"},"PeriodicalIF":1.6,"publicationDate":"2018-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.5511/PLANTBIOTECHNOLOGY.18.0510A","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41555697","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-09-25DOI: 10.5511/PLANTBIOTECHNOLOGY.18.0525A
Seung-Won Choi, Ken Hoshikawa, S. Fujita, D. Thi, T. Mizoguchi, H. Ezura, Emi Ito
Quantitative real-time PCR (qRT-PCR) is widely used to analyze the expression profiles of the genes of interest. In order to obtain accurate quantification data, normalization by using reliable internal control genes is essential. In this study, we evaluated the stability and applicability of eight internal control gene candidates for analyzing gene expression during fruit development in dwarf tomato cultivar Micro-Tom. We collected seventeen different samples from flowers and fruits at different developmental stages, and estimated the expression stability of the candidate genes by two statistical algorithms, geNorm and NormFinder. The combined ranking order and qRT-PCR analyses for expression profiles of SlYABBY2a, SlYABBY1a, FRUITFULL1 and APETALA2c suggested that EXPRESSED was the most stable and reliable internal control gene among the candidates. Our analysis also suggested that RPL8 was also suitable if the sample group is limited to fruits at different maturation stages. In addition to EXPRESSED, GAPDH was also applicable for relative quantitation to monitor gene expression profiles through fruit development from pistil to pericarp.
{"title":"Evaluation of internal control genes for quantitative realtime PCR analyses for studying fruit development of dwarf tomato cultivar 'Micro-Tom'.","authors":"Seung-Won Choi, Ken Hoshikawa, S. Fujita, D. Thi, T. Mizoguchi, H. Ezura, Emi Ito","doi":"10.5511/PLANTBIOTECHNOLOGY.18.0525A","DOIUrl":"https://doi.org/10.5511/PLANTBIOTECHNOLOGY.18.0525A","url":null,"abstract":"Quantitative real-time PCR (qRT-PCR) is widely used to analyze the expression profiles of the genes of interest. In order to obtain accurate quantification data, normalization by using reliable internal control genes is essential. In this study, we evaluated the stability and applicability of eight internal control gene candidates for analyzing gene expression during fruit development in dwarf tomato cultivar Micro-Tom. We collected seventeen different samples from flowers and fruits at different developmental stages, and estimated the expression stability of the candidate genes by two statistical algorithms, geNorm and NormFinder. The combined ranking order and qRT-PCR analyses for expression profiles of SlYABBY2a, SlYABBY1a, FRUITFULL1 and APETALA2c suggested that EXPRESSED was the most stable and reliable internal control gene among the candidates. Our analysis also suggested that RPL8 was also suitable if the sample group is limited to fruits at different maturation stages. In addition to EXPRESSED, GAPDH was also applicable for relative quantitation to monitor gene expression profiles through fruit development from pistil to pericarp.","PeriodicalId":20411,"journal":{"name":"Plant Biotechnology","volume":"35 3 1","pages":"225-235"},"PeriodicalIF":1.6,"publicationDate":"2018-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.5511/PLANTBIOTECHNOLOGY.18.0525A","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"45283655","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-09-25DOI: 10.5511/PLANTBIOTECHNOLOGY.18.0501A
Hiroyuki Tanaka, R. Suzuki, Nanako Okabe, Tomohiro Suzuki, Y. Kodama
Salinity stress limits plant growth and productivity. To cope with this limitation, the expression patterns of numerous genes are altered in response to salt stress; however, the regulatory mechanisms involved in these changes are unclear. In the present study, we investigated the regulation of the salinity stress response in the liverwort Marchantia polymorpha. The growth of M. polymorpha gemmalings was severely inhibited by NaCl, and RNA-sequencing and quantitative RT-PCR analyses revealed that the expression of several transcription factor gene families was induced by salinity stress. This work provides insight into the molecular mechanisms underlying the salinity stress response in M. polymorpha.
{"title":"Salinity stress-responsive transcription factors in the liverwort Marchantia polymorpha.","authors":"Hiroyuki Tanaka, R. Suzuki, Nanako Okabe, Tomohiro Suzuki, Y. Kodama","doi":"10.5511/PLANTBIOTECHNOLOGY.18.0501A","DOIUrl":"https://doi.org/10.5511/PLANTBIOTECHNOLOGY.18.0501A","url":null,"abstract":"Salinity stress limits plant growth and productivity. To cope with this limitation, the expression patterns of numerous genes are altered in response to salt stress; however, the regulatory mechanisms involved in these changes are unclear. In the present study, we investigated the regulation of the salinity stress response in the liverwort Marchantia polymorpha. The growth of M. polymorpha gemmalings was severely inhibited by NaCl, and RNA-sequencing and quantitative RT-PCR analyses revealed that the expression of several transcription factor gene families was induced by salinity stress. This work provides insight into the molecular mechanisms underlying the salinity stress response in M. polymorpha.","PeriodicalId":20411,"journal":{"name":"Plant Biotechnology","volume":"35 3 1","pages":"281-284"},"PeriodicalIF":1.6,"publicationDate":"2018-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.5511/PLANTBIOTECHNOLOGY.18.0501A","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"43781743","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-09-25DOI: 10.5511/PLANTBIOTECHNOLOGY.18.0510B
N. T. Tran, Taichi Oguchi, E. Matsunaga, A. Kawaoka, Kazuo N. Watanabe, A. Kikuchi
Novel transgenic Eucalyptus camaldulensis trees expressing the bacterial choline oxidase A (codA) gene by the Cauliflower mosaic virus (CaMV) 35S promoter and the Arabidopsis thaliana heat shock protein (HSP) terminator was developed. To evaluate the codA transcription level and the metabolic products and abiotic stress tolerance of the transgenic trees, a six-month semi-confined screen house cultivation trial was conducted under a moderate-stringency salt-stress condition. The transcription level of the CaMV 35S promoter driven-codA was more than fourfold higher, and the content of glycine betaine, the metabolic product of codA, was twofold higher, with the HSP terminator than with the nopaline synthase (NOS) terminator. Moreover, the screen house cultivation revealed that the growth of transgenic trees under the salt stress condition was alleviated in correlation with the glycine betaine concentration. These results suggest that the enhancement of codA transcription by the HSP terminator increased the abiotic stress tolerance of Eucalyptus plantation trees.
{"title":"Transcriptional enhancement of a bacterial choline oxidase A gene by an HSP terminator improves the glycine betaine production and salinity stress tolerance of Eucalyptus camaldulensis trees.","authors":"N. T. Tran, Taichi Oguchi, E. Matsunaga, A. Kawaoka, Kazuo N. Watanabe, A. Kikuchi","doi":"10.5511/PLANTBIOTECHNOLOGY.18.0510B","DOIUrl":"https://doi.org/10.5511/PLANTBIOTECHNOLOGY.18.0510B","url":null,"abstract":"Novel transgenic Eucalyptus camaldulensis trees expressing the bacterial choline oxidase A (codA) gene by the Cauliflower mosaic virus (CaMV) 35S promoter and the Arabidopsis thaliana heat shock protein (HSP) terminator was developed. To evaluate the codA transcription level and the metabolic products and abiotic stress tolerance of the transgenic trees, a six-month semi-confined screen house cultivation trial was conducted under a moderate-stringency salt-stress condition. The transcription level of the CaMV 35S promoter driven-codA was more than fourfold higher, and the content of glycine betaine, the metabolic product of codA, was twofold higher, with the HSP terminator than with the nopaline synthase (NOS) terminator. Moreover, the screen house cultivation revealed that the growth of transgenic trees under the salt stress condition was alleviated in correlation with the glycine betaine concentration. These results suggest that the enhancement of codA transcription by the HSP terminator increased the abiotic stress tolerance of Eucalyptus plantation trees.","PeriodicalId":20411,"journal":{"name":"Plant Biotechnology","volume":"35 3 1","pages":"215-224"},"PeriodicalIF":1.6,"publicationDate":"2018-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.5511/PLANTBIOTECHNOLOGY.18.0510B","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"48099410","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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