Detection of Green Fluorescent Protein in a Single Bacterium by Capillary Electrophoresis with Laser-Induced Fluorescence
Green fluorescence protein (GFP) is a common reporter used to monitor protein expression in single cells. However, autofluorescence from endogenous components can mask the signal from GFP, particularly at low expression levels in prokaryotes. We employ capillary electrophoresis with laser-induced fl...
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| Veröffentlicht in: | Analytical chemistry (Washington) 2007-01, Vol.79 (2), p.778-781 |
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| creator | Turner, Emily H Lauterbach, Kevin Pugsley, Haley R Palmer, Vanessa R Dovichi, Norman J |
| description | Green fluorescence protein (GFP) is a common reporter used to monitor protein expression in single cells. However, autofluorescence from endogenous components can mask the signal from GFP, particularly at low expression levels in prokaryotes. We employ capillary electrophoresis with laser-induced fluorescence for the analysis of the expression of green fluorescent protein in a single bacterium. Capillary electrophoresis separates GFP from native cellular autofluorescent components, reducing the background signal and improving detection limits. Our system provides 100 ymol (60 copies) limits of detection for GFP. To demonstrate the performance of this instrument, we employ a model system of Deinococcus radiodurans that has been engineered to express GFP under the control of the recA promoter. We report resolution and detection of GFP and autofluorescent components in a single D. radiodurans bacterium. This paper presents the first example of expression of GFP in D. radiodurans and the first detection of GFP in a single bacterium by capillary electrophoresis. |
| doi_str_mv | 10.1021/ac061778r |
| format | Article |
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Chem</addtitle><description>Green fluorescence protein (GFP) is a common reporter used to monitor protein expression in single cells. However, autofluorescence from endogenous components can mask the signal from GFP, particularly at low expression levels in prokaryotes. We employ capillary electrophoresis with laser-induced fluorescence for the analysis of the expression of green fluorescent protein in a single bacterium. Capillary electrophoresis separates GFP from native cellular autofluorescent components, reducing the background signal and improving detection limits. Our system provides 100 ymol (60 copies) limits of detection for GFP. To demonstrate the performance of this instrument, we employ a model system of Deinococcus radiodurans that has been engineered to express GFP under the control of the recA promoter. We report resolution and detection of GFP and autofluorescent components in a single D. radiodurans bacterium. 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Lauterbach, Kevin ; Pugsley, Haley R ; Palmer, Vanessa R ; Dovichi, Norman J</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a408t-c79e5ffa1df62e490ef36ee99dcf09dc172fb4b01cefffac0896bc479c9dee333</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2007</creationdate><topic>Analytical chemistry</topic><topic>Bacteria</topic><topic>Chemistry</topic><topic>Chromatographic methods and physical methods associated with chromatography</topic><topic>Deinococcus - chemistry</topic><topic>Deinococcus - genetics</topic><topic>Deinococcus radiodurans</topic><topic>Electrophoresis, Capillary - methods</topic><topic>Exact sciences and technology</topic><topic>Fluorescence</topic><topic>Gene expression</topic><topic>General, instrumentation</topic><topic>Green Fluorescent Proteins - analysis</topic><topic>Green Fluorescent Proteins - genetics</topic><topic>Lasers</topic><topic>Other chromatographic methods</topic><topic>Proteins</topic><topic>Scientific apparatus & instruments</topic><topic>Spectrometric and optical methods</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Turner, Emily H</creatorcontrib><creatorcontrib>Lauterbach, Kevin</creatorcontrib><creatorcontrib>Pugsley, Haley R</creatorcontrib><creatorcontrib>Palmer, Vanessa R</creatorcontrib><creatorcontrib>Dovichi, Norman J</creatorcontrib><collection>Istex</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Aluminium Industry Abstracts</collection><collection>Biotechnology Research Abstracts</collection><collection>Ceramic Abstracts</collection><collection>Computer and Information Systems Abstracts</collection><collection>Corrosion Abstracts</collection><collection>Electronics & Communications Abstracts</collection><collection>Engineered Materials Abstracts</collection><collection>Materials Business File</collection><collection>Mechanical & Transportation Engineering Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>Toxicology Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>METADEX</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ANTE: Abstracts in New Technology & Engineering</collection><collection>Engineering Research Database</collection><collection>Aerospace Database</collection><collection>Copper Technical Reference Library</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Materials Research Database</collection><collection>ProQuest Computer Science Collection</collection><collection>Civil Engineering Abstracts</collection><collection>Advanced Technologies Database with Aerospace</collection><collection>Computer and Information Systems Abstracts Academic</collection><collection>Computer and Information Systems Abstracts Professional</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><jtitle>Analytical chemistry (Washington)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Turner, Emily H</au><au>Lauterbach, Kevin</au><au>Pugsley, Haley R</au><au>Palmer, Vanessa R</au><au>Dovichi, Norman J</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Detection of Green Fluorescent Protein in a Single Bacterium by Capillary Electrophoresis with Laser-Induced Fluorescence</atitle><jtitle>Analytical chemistry (Washington)</jtitle><addtitle>Anal. Chem</addtitle><date>2007-01-15</date><risdate>2007</risdate><volume>79</volume><issue>2</issue><spage>778</spage><epage>781</epage><pages>778-781</pages><issn>0003-2700</issn><eissn>1520-6882</eissn><coden>ANCHAM</coden><abstract>Green fluorescence protein (GFP) is a common reporter used to monitor protein expression in single cells. However, autofluorescence from endogenous components can mask the signal from GFP, particularly at low expression levels in prokaryotes. We employ capillary electrophoresis with laser-induced fluorescence for the analysis of the expression of green fluorescent protein in a single bacterium. Capillary electrophoresis separates GFP from native cellular autofluorescent components, reducing the background signal and improving detection limits. Our system provides 100 ymol (60 copies) limits of detection for GFP. To demonstrate the performance of this instrument, we employ a model system of Deinococcus radiodurans that has been engineered to express GFP under the control of the recA promoter. We report resolution and detection of GFP and autofluorescent components in a single D. radiodurans bacterium. This paper presents the first example of expression of GFP in D. radiodurans and the first detection of GFP in a single bacterium by capillary electrophoresis.</abstract><cop>Washington, DC</cop><pub>American Chemical Society</pub><pmid>17222051</pmid><doi>10.1021/ac061778r</doi><tpages>4</tpages></addata></record> |
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| subjects | Analytical chemistry Bacteria Chemistry Chromatographic methods and physical methods associated with chromatography Deinococcus - chemistry Deinococcus - genetics Deinococcus radiodurans Electrophoresis, Capillary - methods Exact sciences and technology Fluorescence Gene expression General, instrumentation Green Fluorescent Proteins - analysis Green Fluorescent Proteins - genetics Lasers Other chromatographic methods Proteins Scientific apparatus & instruments Spectrometric and optical methods |
| title | Detection of Green Fluorescent Protein in a Single Bacterium by Capillary Electrophoresis with Laser-Induced Fluorescence |
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