A rapid, highly efficient and economical method of Agrobacterium-mediated in planta transient transformation in living onion epidermis
Transient transformation is simpler, more efficient and economical in analyzing protein subcellular localization than stable transformation. Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living an...
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| Veröffentlicht in: | PloS one 2014-01, Vol.9 (1), p.e83556-e83556 |
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| creator | Xu, Kedong Huang, Xiaohui Wu, Manman Wang, Yan Chang, Yunxia Liu, Kun Zhang, Ju Zhang, Yi Zhang, Fuli Yi, Liming Li, Tingting Wang, Ruiyue Tan, Guangxuan Li, Chengwei |
| description | Transient transformation is simpler, more efficient and economical in analyzing protein subcellular localization than stable transformation. Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale. |
| doi_str_mv | 10.1371/journal.pone.0083556 |
| format | Article |
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Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0083556</identifier><identifier>PMID: 24416168</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Agriculture ; Agrobacterium - metabolism ; Arabidopsis ; Arabidopsis - microbiology ; Biolistics ; Biology ; Bombardment ; Economic analysis ; Efficiency ; Epidermis ; Equipment costs ; Fluorescence ; Gene expression ; Genetic Techniques - economics ; Genetic transformation ; Kinases ; Laboratories ; Life sciences ; Localization ; Methods ; Monomolecular films ; Nicotiana - microbiology ; Onions - genetics ; Onions - microbiology ; Oxidative stress ; Panicum virgatum ; Particle bombardment ; Phase transitions ; Plant cells ; Plant Epidermis - cytology ; Plant Epidermis - microbiology ; Plants (botany) ; Plants, Genetically Modified ; Populus euphratica ; Proteins ; Reproducibility of Results ; Subcellular Fractions - metabolism ; Time Factors ; Tobacco ; Transfection ; Transformation ; Transformation, Genetic ; Vegetables</subject><ispartof>PloS one, 2014-01, Vol.9 (1), p.e83556-e83556</ispartof><rights>COPYRIGHT 2014 Public Library of Science</rights><rights>2014 Xu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2014 Xu et al 2014 Xu et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c692t-c88f98bf5b40a35e7edc54430ef1e7611f26f8d34ccc998489884a1b91308fcb3</citedby><cites>FETCH-LOGICAL-c692t-c88f98bf5b40a35e7edc54430ef1e7611f26f8d34ccc998489884a1b91308fcb3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3885512/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3885512/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,860,881,2095,2914,23846,27903,27904,53770,53772,79347,79348</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24416168$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Davis, Keith R.</contributor><creatorcontrib>Xu, Kedong</creatorcontrib><creatorcontrib>Huang, Xiaohui</creatorcontrib><creatorcontrib>Wu, Manman</creatorcontrib><creatorcontrib>Wang, Yan</creatorcontrib><creatorcontrib>Chang, Yunxia</creatorcontrib><creatorcontrib>Liu, Kun</creatorcontrib><creatorcontrib>Zhang, Ju</creatorcontrib><creatorcontrib>Zhang, Yi</creatorcontrib><creatorcontrib>Zhang, Fuli</creatorcontrib><creatorcontrib>Yi, Liming</creatorcontrib><creatorcontrib>Li, Tingting</creatorcontrib><creatorcontrib>Wang, Ruiyue</creatorcontrib><creatorcontrib>Tan, Guangxuan</creatorcontrib><creatorcontrib>Li, Chengwei</creatorcontrib><title>A rapid, highly efficient and economical method of Agrobacterium-mediated in planta transient transformation in living onion epidermis</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>Transient transformation is simpler, more efficient and economical in analyzing protein subcellular localization than stable transformation. Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale.</description><subject>Agriculture</subject><subject>Agrobacterium - metabolism</subject><subject>Arabidopsis</subject><subject>Arabidopsis - microbiology</subject><subject>Biolistics</subject><subject>Biology</subject><subject>Bombardment</subject><subject>Economic analysis</subject><subject>Efficiency</subject><subject>Epidermis</subject><subject>Equipment costs</subject><subject>Fluorescence</subject><subject>Gene expression</subject><subject>Genetic Techniques - economics</subject><subject>Genetic transformation</subject><subject>Kinases</subject><subject>Laboratories</subject><subject>Life sciences</subject><subject>Localization</subject><subject>Methods</subject><subject>Monomolecular films</subject><subject>Nicotiana - microbiology</subject><subject>Onions - genetics</subject><subject>Onions - microbiology</subject><subject>Oxidative stress</subject><subject>Panicum virgatum</subject><subject>Particle bombardment</subject><subject>Phase transitions</subject><subject>Plant cells</subject><subject>Plant Epidermis - cytology</subject><subject>Plant Epidermis - microbiology</subject><subject>Plants (botany)</subject><subject>Plants, Genetically Modified</subject><subject>Populus euphratica</subject><subject>Proteins</subject><subject>Reproducibility of Results</subject><subject>Subcellular Fractions - metabolism</subject><subject>Time Factors</subject><subject>Tobacco</subject><subject>Transfection</subject><subject>Transformation</subject><subject>Transformation, Genetic</subject><subject>Vegetables</subject><issn>1932-6203</issn><issn>1932-6203</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>DOA</sourceid><recordid>eNqNk12L1DAUhoso7rr6D0QLgig4Y9KkaXojDIsfAwsLft2GNB9tljYZk3Rx_4C_23Smu0xlL6QXTdPnfU_z9pwsew7BGqIKvr9yo7e8X--cVWsAKCpL8iA7hTUqVqQA6OHR-iR7EsIVACWihDzOTgqMIYGEnmZ_NrnnOyPf5Z1pu_4mV1obYZSNObcyV8JZNxjB-3xQsXMydzrftN41XETlzTisBiUNj0rmxua7ntvI8-i5DXuP_Uo7P_BonJ2Q3lwb2-bOTs8qVVZ-MOFp9kjzPqhn8_0s-_Hp4_fzL6uLy8_b883FSpC6iCtBqa5po8sGA45KVSkpSowRUBqqikCoC6KpRFgIUdcU05pSzGFTQwSoFg06y14efHe9C2yOMDCIk5gCgEgitgdCOn7Fdt4M3N8wxw3bbzjfMu6jEb1iddOIZEoLVCDMJWmElJUglCsoIWmK5PVhrjY2KSaREvG8X5gu31jTsdZdM0RpWcLJ4M1s4N2vUYXIUlZC9Slm5cbpu2tQEVxCmtBX_6D3n26mWp4OYKx2qa6YTNkGV5RWGFZT2fU9VLqkSr2Q-k2btL8QvF0IEhPV79jyMQS2_fb1_9nLn0v29RHbKd7HLrh-nJopLEF8AIV3IXil70KGgE3jcpsGm8aFzeOSZC-Of9Cd6HY-0F_oBhLM</recordid><startdate>20140108</startdate><enddate>20140108</enddate><creator>Xu, Kedong</creator><creator>Huang, Xiaohui</creator><creator>Wu, Manman</creator><creator>Wang, Yan</creator><creator>Chang, Yunxia</creator><creator>Liu, Kun</creator><creator>Zhang, Ju</creator><creator>Zhang, Yi</creator><creator>Zhang, Fuli</creator><creator>Yi, Liming</creator><creator>Li, Tingting</creator><creator>Wang, Ruiyue</creator><creator>Tan, Guangxuan</creator><creator>Li, Chengwei</creator><general>Public Library of Science</general><general>Public Library of Science (PLoS)</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>IOV</scope><scope>ISR</scope><scope>3V.</scope><scope>7QG</scope><scope>7QL</scope><scope>7QO</scope><scope>7RV</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TG</scope><scope>7TM</scope><scope>7U9</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>KL.</scope><scope>L6V</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>M7S</scope><scope>NAPCQ</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PATMY</scope><scope>PDBOC</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PTHSS</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope><scope>DOA</scope></search><sort><creationdate>20140108</creationdate><title>A rapid, highly efficient and economical method of Agrobacterium-mediated in planta transient transformation in living onion epidermis</title><author>Xu, Kedong ; Huang, Xiaohui ; Wu, Manman ; Wang, Yan ; Chang, Yunxia ; Liu, Kun ; Zhang, Ju ; Zhang, Yi ; Zhang, Fuli ; Yi, Liming ; Li, Tingting ; Wang, Ruiyue ; Tan, Guangxuan ; Li, Chengwei</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c692t-c88f98bf5b40a35e7edc54430ef1e7611f26f8d34ccc998489884a1b91308fcb3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2014</creationdate><topic>Agriculture</topic><topic>Agrobacterium - 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Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>24416168</pmid><doi>10.1371/journal.pone.0083556</doi><tpages>e83556</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | PloS one, 2014-01, Vol.9 (1), p.e83556-e83556 |
| issn | 1932-6203 1932-6203 |
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| source | MEDLINE; DOAJ Directory of Open Access Journals; Public Library of Science (PLoS) Journals Open Access; EZB-FREE-00999 freely available EZB journals; PubMed Central; Free Full-Text Journals in Chemistry |
| subjects | Agriculture Agrobacterium - metabolism Arabidopsis Arabidopsis - microbiology Biolistics Biology Bombardment Economic analysis Efficiency Epidermis Equipment costs Fluorescence Gene expression Genetic Techniques - economics Genetic transformation Kinases Laboratories Life sciences Localization Methods Monomolecular films Nicotiana - microbiology Onions - genetics Onions - microbiology Oxidative stress Panicum virgatum Particle bombardment Phase transitions Plant cells Plant Epidermis - cytology Plant Epidermis - microbiology Plants (botany) Plants, Genetically Modified Populus euphratica Proteins Reproducibility of Results Subcellular Fractions - metabolism Time Factors Tobacco Transfection Transformation Transformation, Genetic Vegetables |
| title | A rapid, highly efficient and economical method of Agrobacterium-mediated in planta transient transformation in living onion epidermis |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-22T06%3A57%3A19IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_plos_&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=A%20rapid,%20highly%20efficient%20and%20economical%20method%20of%20Agrobacterium-mediated%20in%20planta%20transient%20transformation%20in%20living%20onion%20epidermis&rft.jtitle=PloS%20one&rft.au=Xu,%20Kedong&rft.date=2014-01-08&rft.volume=9&rft.issue=1&rft.spage=e83556&rft.epage=e83556&rft.pages=e83556-e83556&rft.issn=1932-6203&rft.eissn=1932-6203&rft_id=info:doi/10.1371/journal.pone.0083556&rft_dat=%3Cgale_plos_%3EA478874172%3C/gale_plos_%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=1476180036&rft_id=info:pmid/24416168&rft_galeid=A478874172&rft_doaj_id=oai_doaj_org_article_9bbcfcb823234ad6bcdd7c68ae1d16b2&rfr_iscdi=true |