François Berthold , David Roujol , Caroline Hemmer , Elisabeth Jamet , Christophe Ritzenthaler , Laurent Hoffmann , Corinne Schmitt-Keichinger
{"title":"里面还是外面?允许植物蛋白亚细胞定位或过度表达的Gateway载体的新集合。","authors":"François Berthold , David Roujol , Caroline Hemmer , Elisabeth Jamet , Christophe Ritzenthaler , Laurent Hoffmann , Corinne Schmitt-Keichinger","doi":"10.1016/j.plasmid.2019.102436","DOIUrl":null,"url":null,"abstract":"<div><p><span><span><span>Transient expression of proteins based on agro-infiltration techniques has proven very efficient and straightforward to study the intrinsic properties of proteins. The </span>level of protein expression has been enhanced by the use of </span>vector plasmids<span> containing virus-derived sequences and the cloning step has been facilitated by recombination technologies. The pEAQ-</span></span><em>HT</em><span>-DEST series of vectors fulfilling these improvements are vectors of choice. However, they lack the possibility to directly and easily fuse the protein of interest to a fluorescent tag<span> or to address it to the secretion pathway. In the present work we describe the production of 15 pEAQ-</span></span><em>HT</em><span>-DEST1-based plasmids designed to use the Gateway® cloning technology and to generate high levels of fluorescent fusion protein by agro-infiltration, </span><em>in planta</em><span><span><span>. This collection of plasmids includes binary vectors allowing N-terminal or C-terminal fusion to the bright tags EGFP or TagRFP for cytoplasmic accumulation or secretion and represents therefore a valuable tool for subcellular localization or biochemical studies. A </span>viral protein, the blue fluorescent protein TagBFP, the </span>green fluorescent protein variant T-Sapphire and an </span><span><em>Arabidopsis</em></span> protein were transiently expressed in <em>N. benthamiana</em> to demonstrate the potential of these vectors.</p></div>","PeriodicalId":49689,"journal":{"name":"Plasmid","volume":null,"pages":null},"PeriodicalIF":1.8000,"publicationDate":"2019-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.plasmid.2019.102436","citationCount":"9","resultStr":"{\"title\":\"Inside or outside? A new collection of Gateway vectors allowing plant protein subcellular localization or over-expression\",\"authors\":\"François Berthold , David Roujol , Caroline Hemmer , Elisabeth Jamet , Christophe Ritzenthaler , Laurent Hoffmann , Corinne Schmitt-Keichinger\",\"doi\":\"10.1016/j.plasmid.2019.102436\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p><span><span><span>Transient expression of proteins based on agro-infiltration techniques has proven very efficient and straightforward to study the intrinsic properties of proteins. The </span>level of protein expression has been enhanced by the use of </span>vector plasmids<span> containing virus-derived sequences and the cloning step has been facilitated by recombination technologies. The pEAQ-</span></span><em>HT</em><span>-DEST series of vectors fulfilling these improvements are vectors of choice. However, they lack the possibility to directly and easily fuse the protein of interest to a fluorescent tag<span> or to address it to the secretion pathway. In the present work we describe the production of 15 pEAQ-</span></span><em>HT</em><span>-DEST1-based plasmids designed to use the Gateway® cloning technology and to generate high levels of fluorescent fusion protein by agro-infiltration, </span><em>in planta</em><span><span><span>. This collection of plasmids includes binary vectors allowing N-terminal or C-terminal fusion to the bright tags EGFP or TagRFP for cytoplasmic accumulation or secretion and represents therefore a valuable tool for subcellular localization or biochemical studies. A </span>viral protein, the blue fluorescent protein TagBFP, the </span>green fluorescent protein variant T-Sapphire and an </span><span><em>Arabidopsis</em></span> protein were transiently expressed in <em>N. benthamiana</em> to demonstrate the potential of these vectors.</p></div>\",\"PeriodicalId\":49689,\"journal\":{\"name\":\"Plasmid\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.8000,\"publicationDate\":\"2019-09-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1016/j.plasmid.2019.102436\",\"citationCount\":\"9\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Plasmid\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0147619X19300289\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"GENETICS & HEREDITY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Plasmid","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0147619X19300289","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"GENETICS & HEREDITY","Score":null,"Total":0}
Inside or outside? A new collection of Gateway vectors allowing plant protein subcellular localization or over-expression
Transient expression of proteins based on agro-infiltration techniques has proven very efficient and straightforward to study the intrinsic properties of proteins. The level of protein expression has been enhanced by the use of vector plasmids containing virus-derived sequences and the cloning step has been facilitated by recombination technologies. The pEAQ-HT-DEST series of vectors fulfilling these improvements are vectors of choice. However, they lack the possibility to directly and easily fuse the protein of interest to a fluorescent tag or to address it to the secretion pathway. In the present work we describe the production of 15 pEAQ-HT-DEST1-based plasmids designed to use the Gateway® cloning technology and to generate high levels of fluorescent fusion protein by agro-infiltration, in planta. This collection of plasmids includes binary vectors allowing N-terminal or C-terminal fusion to the bright tags EGFP or TagRFP for cytoplasmic accumulation or secretion and represents therefore a valuable tool for subcellular localization or biochemical studies. A viral protein, the blue fluorescent protein TagBFP, the green fluorescent protein variant T-Sapphire and an Arabidopsis protein were transiently expressed in N. benthamiana to demonstrate the potential of these vectors.
期刊介绍:
Plasmid publishes original research on genetic elements in all kingdoms of life with emphasis on maintenance, transmission and evolution of extrachromosomal elements. Objects of interest include plasmids, bacteriophages, mobile genetic elements, organelle DNA, and genomic and pathogenicity islands.