Difference between revisions of "Team:Alverno CA/Team"

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{{Alverno_CA}}
 
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    rel="icon"><br>
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    <br>
  
  
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    <center>
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        <img alt="Alverno iGEM Logo" src=
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        "https://static.igem.org/mediawiki/2016/thumb/5/58/T--Alverno_CA--Alverno_iGEM_2016_Logo.png/600px-T--Alverno_CA--Alverno_iGEM_2016_Logo.png"
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<head>
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<br><br>
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        <h2>Interlab Study</h2>
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<title>Alverno iGEM 2016</title>
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    </center>
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<link rel="icon" href="https://static.igem.org/mediawiki/2016/thumb/5/58/T--Alverno_CA--Alverno_iGEM_2016_Logo.png/600px-T--Alverno_CA--Alverno_iGEM_2016_Logo.png">
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<center><img src="https://static.igem.org/mediawiki/2016/thumb/5/58/T--Alverno_CA--Alverno_iGEM_2016_Logo.png/600px-T--Alverno_CA--Alverno_iGEM_2016_Logo.png" alt="Alverno iGEM Logo" style="width:300px;"></center>
 
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<center><h2>Our Team</h2></center>
 
  
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         <div class="demo" id="container">
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         <h3>Quantifying different sources of variation in gene expression<br> using
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        fluorescent transformed E. coli bacteria</h3>
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    </center>
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    <div class="demo" id="container">
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                 <img style="width:925px;margin-top:146px;" src="https://static.igem.org/mediawiki/2016/4/48/T--Alverno_CA--TeamGroup.jpg"  title="We're the Alverno_CA iGEM Team!"/>
 
                 <img style="width:925px;margin-top:146px;" src="https://static.igem.org/mediawiki/2016/4/48/T--Alverno_CA--TeamGroup.jpg"  title="We're the Alverno_CA iGEM Team!"/>
 
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I spend my life laughing about band memes and and watching YouTube.<br><br>HS Freshman; Member of Alverno's Accelerated Honors Academy for Gifted Girls; Science Best in Class (8th grade)"/>
 
I spend my life laughing about band memes and and watching YouTube.<br><br>HS Freshman; Member of Alverno's Accelerated Honors Academy for Gifted Girls; Science Best in Class (8th grade)"/>
 
    </li>               
 
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    <br>
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 +
 +
    <h5>
 +
    </h5>
 +
 +
 +
    <h4>Summary:</h4>
 +
    <h5></h5>
 +
 +
    <h5>As part of the 2016 iGEM Interlab Study, we tested 3 different
 +
    fluorescent protein expression plasmids, J23101, J23106, and J23117 with
 +
    positive and negative controls. We transformed the parts with iGEM's
 +
    transformation protocol. Our bacteria were cultured on a LB agar plate and
 +
    cultured in LB broth. To measure the differences in fluorescence expressed
 +
    by different strength plasmids, we used a plate reader with an OD600
 +
    reference point and FITC standard curve.</h5>
 +
 +
 +
    <h5>
 +
    </h5>
 +
<br><br>
 +
 +
    <h4>Project Description:</h4>
 +
    <h5></h5>
 +
 +
    <h5>The 2016 iGEM Interlab Study aims to compare results obtained by various
 +
    teams in order to quantify the expression of 5 different constructs using
 +
    fluorescence, which provides a useful insight into expression levels that
 +
    can be monitored without disrupting cells. Each device used in the study is
 +
    in the pSB1C3 plasmid backbone and are fluorescent protein expression
 +
    plasmids of different strength promoters. The devices we tested were
 +
    J23101, J23106, J23117 (Test Devices 1, 2, and 3). We used positive control
 +
    I20270 and negative control R0040. J23101, with a strong promoter, showed
 +
    the highest amount of fluorescence. J23117, with a weak promoter, showed
 +
    the lowest amount of fluorescence, and J23106, with the medium strength
 +
    promoter, showed fluorescence amounts between the other two. Our test
 +
    devices were inserted into DH5alpha E. coli, which were transformed
 +
    according to the iGEM transformation protocol. Our transformed cells were
 +
    plated on LB plates with chloramphenicol and incubated overnight at 37° C.
 +
    Two colonies were picked from each plate (with the exception of Test Device
 +
    3, with which there was only one) and were inoculated in 15mL test tubes.
 +
    These devices were diluted, and then measured in a plate reader with an
 +
    excitation wavelength of 35nm, according to iGEM’s measurement
 +
    protocol.</h5>
 +
 +
 +
    <h5>
 +
    </h5>
 +
 +
 +
    <h5>
 +
    </h5>
 +
<br><br>
 +
 +
    <h4>Protocol:</h4>
 +
    <h5></h5>
 +
 +
    <h5><a href=
 +
    "https://static.igem.org/mediawiki/2016/c/c5/InterLab_iGEM2016_Plate_Reader_Protocol_Updated_July.pdf">
 +
    Plate Reader Protocol</a>
 +
    </h5>
 +
 +
 +
    <h5><a href=
 +
    "https://static.igem.org/mediawiki/parts/6/67/IGEM_Registry_-_Transformation_Protocol.pdf">
 +
    Transformation Protocol</a>
 +
    </h5>
 +
 +
 +
    <h5>
 +
    </h5>
 +
 +
<br><br>
 +
    <h4>Results:</h4>
 +
    <h5></h5>
 +
 +
    <center>
 +
        <center>
 +
            <table class="image">
 +
                <tr>
 +
                    <td><img alt="Placeholder Image" src=
 +
                    "http://i.imgur.com/FcVrzL0.png" style="width:800px;">
 +
                    </td>
 +
                </tr>
 +
 +
 +
                <tr>
 +
                    <td class="caption">Graph of Absorbance (600) over a 6 hour
 +
                    time period</td>
 +
                </tr>
 +
            </table>
 +
        </center>
 +
 +
 +
        <center>
 +
            <table class="image">
 +
                <tr>
 +
                    <td><img alt="Placeholder Image" src=
 +
                    "http://i.imgur.com/eKgjQDe.png" style="width:800px;">
 +
                    </td>
 +
                </tr>
 +
 +
 +
                <tr>
 +
                    <td class="caption">Graph of Fluorescence in AU over a 6
 +
                    hour time period</td>
 +
                </tr>
 +
            </table>
 +
        </center>
 +
 +
 +
        <center>
 +
            <table class="image">
 +
                <tr>
 +
                    <td><img alt="Placeholder Image" src=
 +
                    "http://i.imgur.com/oyXnJRE.png" style="width:800px;">
 +
                    </td>
 +
                </tr>
 +
 +
 +
                <tr>
 +
                    <td class="caption">Graph of Fluorescence/Absorbance in AU
 +
                    over a 6 hour time period</td>
 +
                </tr>
 +
            </table>
 +
        </center>
 +
 +
 +
        <table class="image">
 +
            <tr>
 +
                <td><img alt="Placeholder Image" src=
 +
                "http://i.imgur.com/lWwA42t.png" style="width:800px;">
 +
                </td>
 +
            </tr>
  
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−
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−
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 +
            <tr>
 +
                <td class="caption">FITC Fluorescence standard curve. Row A had
 +
                pipetting errors and so is different from the other
 +
                curves.</td>
 +
            </tr>
 +
        </table>
 +
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Revision as of 00:19, 19 October 2016

Alverno iGEM 2016

Alverno iGEM Logo

Interlab Study

Quantifying different sources of variation in gene expression
using fluorescent transformed E. coli bacteria





























Summary:

As part of the 2016 iGEM Interlab Study, we tested 3 different fluorescent protein expression plasmids, J23101, J23106, and J23117 with positive and negative controls. We transformed the parts with iGEM's transformation protocol. Our bacteria were cultured on a LB agar plate and cultured in LB broth. To measure the differences in fluorescence expressed by different strength plasmids, we used a plate reader with an OD600 reference point and FITC standard curve.


Project Description:

The 2016 iGEM Interlab Study aims to compare results obtained by various teams in order to quantify the expression of 5 different constructs using fluorescence, which provides a useful insight into expression levels that can be monitored without disrupting cells. Each device used in the study is in the pSB1C3 plasmid backbone and are fluorescent protein expression plasmids of different strength promoters. The devices we tested were J23101, J23106, J23117 (Test Devices 1, 2, and 3). We used positive control I20270 and negative control R0040. J23101, with a strong promoter, showed the highest amount of fluorescence. J23117, with a weak promoter, showed the lowest amount of fluorescence, and J23106, with the medium strength promoter, showed fluorescence amounts between the other two. Our test devices were inserted into DH5alpha E. coli, which were transformed according to the iGEM transformation protocol. Our transformed cells were plated on LB plates with chloramphenicol and incubated overnight at 37° C. Two colonies were picked from each plate (with the exception of Test Device 3, with which there was only one) and were inoculated in 15mL test tubes. These devices were diluted, and then measured in a plate reader with an excitation wavelength of 35nm, according to iGEM’s measurement protocol.


Protocol:

Plate Reader Protocol
Transformation Protocol


Results:

Placeholder Image
Graph of Absorbance (600) over a 6 hour time period
Placeholder Image
Graph of Fluorescence in AU over a 6 hour time period
Placeholder Image
Graph of Fluorescence/Absorbance in AU over a 6 hour time period
Placeholder Image
FITC Fluorescence standard curve. Row A had pipetting errors and so is different from the other curves.