Difference between revisions of "Team:Tokyo Tech/InterLab"

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  <!-- Site made with Mobirise Website Builder v4.8.4, https://mobirise.com -->
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  <title>InterLab</title>
  
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<h3>★  ALERT! </h3>
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  <link rel="stylesheet" href="https://2018.igem.org/Team:Tokyo_Tech/mobirise/mobirise/css/mbr-additional-css-interlab?action=raw&amp;ctype=text/css">
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<p>This page is used by the judges to evaluate your team for the <a href="https://2018.igem.org/Judging/Medals">medal criterion</a> or <a href="https://2018.igem.org/Judging/Awards"> award listed below</a>. </p>
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<p> Delete this box in order to be evaluated for this medal criterion and/or award. See more information at <a href="https://2018.igem.org/Judging/Pages_for_Awards"> Instructions for Pages for awards</a>.</p>
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<h1>InterLab</h1>
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<h3>Bronze Medal Criterion #4</h3>
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<p><b>Standard Tracks:</b> Participate in the Interlab Measurement Study and/or obtain new, high quality experimental characterization data for an existing BioBrick Part or Device and enter this information on that part's Main Page in the Registry. The part that you are characterizing must NOT be from a 2018 part number range.
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For teams participating in the <a href="https://2018.igem.org/Measurement/InterLab">InterLab study</a>, all work must be shown on this page.
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<section class="engine"><a href="https://mobirise.info/k">how to develop your own website</a></section><section class="header10 cid-r5UmXXtySa mbr-fullscreen mbr-parallax-background" id="header10-1f">
  
  
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    <div class="container">
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            <h1 class="mbr-section-title align-right mbr-bold pb-3 mbr-fonts-style display-1">InterLab</h1>
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            <h3 class="mbr-text align-right pb-3 mbr-fonts-style display-5">We introduced eight plasmids (2 controls and 6 test devices) to E.coli DH5α by following the protocol and then evaluate the measurement by measuring its OD600. We used TECAN infinite M200 PRO as a plate reader.</h3>
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        </div>
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    </div>
  
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    <div class="mbr-arrow hidden-sm-down" aria-hidden="true">
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        <a href="#next">
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            <i class="mbri-down mbr-iconfont"></i>
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        </a>
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    </div>
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</section>
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<section class="mbr-section article content10 cid-r5UnBb2uc4" id="content10-1g">
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    <div class="container">
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        <div class="inner-container" style="width: 81%;">
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            <hr class="line" style="width: 25%;">
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              <div class="section-text align-center mbr-white mbr-fonts-style display-5">"Not only in the synthetic biology, but in the general scientific field, it is definitely important to get the result with high reproducibility and high credibility."</div><br>
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              <div class="section-text align-center mbr-white mbr-fonts-style display-5">"To achieve this, in iGEM, InterLab Study has been conducted by collecting and comparing the result of GFP measurements with a plate reader."</div><br>
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              <div class="section-text align-center mbr-white mbr-fonts-style display-5"><a href="https://static.igem.org/mediawiki/2018/0/09/2018_InterLab_Plate_Reader_Protocol.pdf" target="_blank">--- iGEM Headquarters</a></div></div>
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            <hr class="line" style="width: 25%;">
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        </div>
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    </div>
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</section>
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<section class="features6 cid-r6rWoqOjeQ" id="features6-1o">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="card p-3 col-12 col-md-6 col-lg-3">
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                <div class="card-img pb-3">
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                    <span class="mbr-iconfont mbri-login"></span>
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                </div>
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                <div class="card-box">
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                    <h1 class="card-title py-3 mbr-fonts-style display-7">Transformation</h1>
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                    <h2 class="mbr-text mbr-fonts-style display-7">We introduced eight plasmids from the Distribution Kit to E.Coli DH5α.&nbsp;</h2>
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                </div>
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            </div>
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            <div class="card p-3 col-12 col-md-6 col-lg-3">
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                <div class="card-img pb-3">
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                    <span class="mbr-iconfont mbri-target"></span>
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                </div>
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                <div class="card-box">
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                    <h3 class="card-title py-3 mbr-fonts-style display-7">Calibration
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</h3>
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                    <h4 class="mbr-text mbr-fonts-style display-7">We conducted three tests and calibrated the machine.</h4>
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                </div>
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            </div>
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            <div class="card p-3 col-12 col-md-6 col-lg-3">
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                <div class="card-img pb-3">
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                    <span class="mbr-iconfont mbri-speed"></span>
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                </div>
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                <div class="card-box">
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                    <h2 class="card-title py-3 mbr-fonts-style display-7">Cell Measurement
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</h2>
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                    <h3 class="mbr-text mbr-fonts-style display-7">We measured the OD600 and the fluorescence of the transformed cells.&nbsp;</h3>
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                </div>
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            </div>
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            <div class="card p-3 col-12 col-md-6 col-lg-3">
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                <div class="card-img pb-3">
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                    <span class="mbri-desktop mbr-iconfont"></span>
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                </div>
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                <div class="card-box">
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                    <h3 class="card-title py-3 mbr-fonts-style display-7">CFU</h3>
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                    <h4 class="mbr-text mbr-fonts-style display-7">
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                      We measured the  Colony Forming Units&nbsp;using two types of transformation cells.</h4>
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                </div>
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            </div>
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</section>
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<section class="testimonials3 cid-r6sb8dQWG4" id="testimonials3-1r">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="media-content px-3 align-self-center mbr-white py-2">
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                <h2 class="mbr-text testimonial-text mbr-fonts-style display-5"><strong>Transformation</strong><br></h2>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">
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                    We took the following eight plasmids from the Distribution Kit and introduced them to E.Coli DH5α. Then, we selected two colonies and cultured them for 16 hours.
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                </span></h3>
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                <p class="mbr-author-desc mbr-fonts-style display-7"><br><br></p>
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            </div>
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            <div class="mbr-figure pl-lg-5" style="width: 140%;">
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              <img src="https://static.igem.org/mediawiki/2018/1/11/T--Tokyo_Tech--Transformation.png" alt="" title="">
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            </div>
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        </div>
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    </div>
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</section>
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<section class="testimonials3 cid-r6sbW6b2I2" id="testimonials3-1s">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="media-content px-3 align-self-center mbr-white py-2">
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                <h2 class="mbr-text testimonial-text mbr-fonts-style display-5"><strong>Calibration</strong></h2>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">We conducted the following three measurements to get the calibration values as a preparation for the Cell Measurement.
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<br></span><br>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">①Measurment of the LUDOX CL-X solution to obtain a conversation factor to transform the  absorbance data into a comparable OD600 measurement.</span></h3>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">②ABS600 measument of the dilution series  of silica beads to construct a standard curve of particle concentration which can be used to convert Abs 600 measurements to an estimated number of cells.&nbsp;</span></h3>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">③Fluorescence measurement of the dilution series of Fluorescein to generate a standard curve of fluorescence for fluorescein concentration. You will be able to use this to convert your cell based readings to an equivalent fluorescein concentration.&nbsp;</span></h3>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;"></span></h3>
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                <p class="mbr-author-desc mbr-fonts-style display-7"><br><br></p>
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            </div>
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            <div class="mbr-figure pl-lg-5" style="width: 130%;">
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              <img src="https://static.igem.org/mediawiki/2018/7/73/T--Tokyo_Tech--Calibration.png" alt="" title="">
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            </div>
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        </div>
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    </div>
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</section>
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<section class="mbr-section article content1 cid-r6skCp9zvU" id="content2-1x">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="mbr-text col-12 col-md-8 mbr-fonts-style display-7">
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                <blockquote><h3><strong>Discussion about Calibration</strong></h3>
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                  <h3>From the Experimental results,Particles / Abs600 = 5.18*10⁸ and MEFL / a.u. = 1.24*10⁹ was found.</h3></blockquote>
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            </div>
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        </div>
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    </div>
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</section>
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<section class="testimonials3 cid-r6sd5F75px" id="testimonials3-1t">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="media-content px-3 align-self-center mbr-white py-2">
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                <h2 class="mbr-text testimonial-text mbr-fonts-style display-5"><strong>
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                  Cell Measurement
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                </strong></h2>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">We measured the OD600 and the fluorescence of the transformed cells in zero and six hours later. Then we evaluated the measurement data by Calibration values.</span>&nbsp;<br></h3>
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            </div>
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            <div class="mbr-figure pl-lg-5" style="width: 60%;">
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              <img src="https://static.igem.org/mediawiki/2018/f/f8/T--Tokyo_Tech--cell_Measurement.png" alt="" title="">
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            </div>
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        </div>
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    </div>
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</section>
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<section class="mbr-section article content1 cid-r6slwJ7zIL" id="content2-1y">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="mbr-text col-12 col-md-8 mbr-fonts-style display-7">
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                <blockquote><h3> <strong>Discussion about Cell Mesurement</strong></h3><h3>The results show the changes in optical density (λ = 600 nm) of the transformed cells. They show that the cells grew steadily.
 +
</h3><h3>However, when we measured the fluorescence, some values appeared negative or go down after 6-hour culturing. Thus, the Fluorescence/OD and MEFL/Particle differed from the theoretical values.
 +
</h3></blockquote>
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            </div>
 +
        </div>
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    </div>
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</section>
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<section class="testimonials3 cid-r6sd6o961y" id="testimonials3-1u">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="media-content px-3 align-self-center mbr-white py-2">
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                <h2 class="mbr-text testimonial-text mbr-fonts-style display-5"><strong>CFU</strong></p>
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                <h3 class="mbr-author-name pt-4 mb-2 mbr-fonts-style display-7"><span style="font-weight: normal;">We diluted both positive and negative controls so that their OD600 became 0.1. Then, we prepared dilution series of them and spread 8.0*10³, 8.0*10⁴, 8.0*10⁵ to the agar medium, counted the number of colonies after 18 hours of cultivation.&nbsp;</span><br></h3>
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            </div>
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            <div class="mbr-figure pl-lg-5" style="width: 35%;">
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              <img src="https://static.igem.org/mediawiki/2018/2/23/T--Tokyo_Tech--CFU.png" alt="" title="">
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            </div>
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        </div>
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    </div>
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</section>
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<section class="mbr-section article content1 cid-r6slZUdqMG" id="content2-1z">
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    <div class="container">
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        <div class="media-container-row">
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            <div class="mbr-text col-12 col-md-8 mbr-fonts-style display-7">
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                <blockquote><h2><strong>Discussion about CFU</strong> </h2><h3>We counted the CFU (Colony Forming Unit) and the following figures show the number of colony on each plate. Then, we calculated the CFU / mL in the starting sample when the optical density (λ = 600 nm) is 0.1.
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</h3></blockquote>
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            </div>
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            <div class="mbr-text col-12 col-md-8 mbr-fonts-style display-5"><h2><strong>Reference
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</strong></h2><p></p><div><strong>　<a href="https://2018.igem.org/Measurement/InterLab">https://2018.igem.org/Measurement/InterLab
 +
</a></strong></div><div><strong>　<a href="https://2017.igem.org/Team:William_and_Mary/Interlab">https://2017.igem.org/Team:William_and_Mary/Interlab
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</a></strong></div><div><strong>
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</strong></div><div><strong><br></strong></div><p></p></div>
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{{Tokyo_Tech/mobirise/footer}}
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{{Tokyo_Tech/mobirise/script}}

Latest revision as of 03:12, 18 October 2018

<!DOCTYPE html>

InterLab
how to develop your own website

InterLab

We introduced eight plasmids (2 controls and 6 test devices) to E.coli DH5α by following the protocol and then evaluate the measurement by measuring its OD600. We used TECAN infinite M200 PRO as a plate reader.

"Not only in the synthetic biology, but in the general scientific field, it is definitely important to get the result with high reproducibility and high credibility."

"To achieve this, in iGEM, InterLab Study has been conducted by collecting and comparing the result of GFP measurements with a plate reader."

--- iGEM Headquarters

Transformation

We introduced eight plasmids from the Distribution Kit to E.Coli DH5α. 

Calibration

We conducted three tests and calibrated the machine.

Cell Measurement

We measured the OD600 and the fluorescence of the transformed cells. 

CFU

We measured the Colony Forming Units using two types of transformation cells.

Transformation

We took the following eight plasmids from the Distribution Kit and introduced them to E.Coli DH5α. Then, we selected two colonies and cultured them for 16 hours.



Calibration

We conducted the following three measurements to get the calibration values as a preparation for the Cell Measurement.

①Measurment of the LUDOX CL-X solution to obtain a conversation factor to transform the absorbance data into a comparable OD600 measurement.

②ABS600 measument of the dilution series of silica beads to construct a standard curve of particle concentration which can be used to convert Abs 600 measurements to an estimated number of cells. 

③Fluorescence measurement of the dilution series of Fluorescein to generate a standard curve of fluorescence for fluorescein concentration. You will be able to use this to convert your cell based readings to an equivalent fluorescein concentration. 



Discussion about Calibration

From the Experimental results,Particles / Abs600 = 5.18*10⁸ and MEFL / a.u. = 1.24*10⁹ was found.

Cell Measurement

We measured the OD600 and the fluorescence of the transformed cells in zero and six hours later. Then we evaluated the measurement data by Calibration values. 

Discussion about Cell Mesurement

The results show the changes in optical density (λ = 600 nm) of the transformed cells. They show that the cells grew steadily.

However, when we measured the fluorescence, some values appeared negative or go down after 6-hour culturing. Thus, the Fluorescence/OD and MEFL/Particle differed from the theoretical values.

CFU

We diluted both positive and negative controls so that their OD600 became 0.1. Then, we prepared dilution series of them and spread 8.0*10³, 8.0*10⁴, 8.0*10⁵ to the agar medium, counted the number of colonies after 18 hours of cultivation. 

Discussion about CFU

We counted the CFU (Colony Forming Unit) and the following figures show the number of colony on each plate. Then, we calculated the CFU / mL in the starting sample when the optical density (λ = 600 nm) is 0.1.

Reference


Address

2 Chome-12-1
Ookayama, Meguro, Tokyo

Contacts

Email: igem2018tokyotech@gmail.com

Links

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