Team:NGSS TR/transformation.html

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<blockquote>
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  <p>&nbsp;</p>
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  <blockquote>
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    <h1>PROTOCOLS</h1>
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    <blockquote>
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      <h2 class="gfdg">6.TRANSFORMATION      </h2>
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      <blockquote>
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        <ol>
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          <li></li>
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          <li>
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            <p>1.Sterilized the enviroment.
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            </p>
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          </li>
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          <li>
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            <p>2.Take the compatent cells from -80 ºC thawing on ice.</p>
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          </li>
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          <li>
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            <p>3.Add 50 µL of thawed competent cells into 1.5ml  tube. </p>
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          </li>
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          <li>
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            <p>4.Add 1 - 2 µL of the resuspended DNA to the 2ml  tube. Pipet up and down a few times, gently. </p>
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          </li>
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          <li>
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            <p>5.Centrifuge  tubes at 3000 rpm for 30-45 seconds.</p>
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          </li>
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          <li>
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            <p>6.Incubate the cells on ice for 45 minutes. </p>
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          </li>
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          <li>
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            <p>7.Heat  the tubes in the 42 C  heat block for a 80 seconds.</p>
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          </li>
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    </ol>
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        <ol start="8" type="1">
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          <li>
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          <p>8.Incubate      the cells on ice for 5 minutes.</p>
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          </li>
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          <li>
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            <p>9.Add 450      μl of LB broth (make sure that the broth does not contain antibiotics and      is not contaminated) to each transformation</p>
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          </li>
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        </ol>
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        <ol><li>
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            <p>10.Incubate the cells at 37ºC for 1 hour while the  tubes are shaking. </p>
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            <p>11.Label petri dish with LB agar and the  appropriate antibiotic. </p>
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          </li>
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          <li>
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            <p>12.Plate 135 µl of the transformation onto the  dishes, and spread. </p>
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          </li>
 +
          <li>
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            <p>13.Incubate the plates at 37ºC for 12-14 hours, making sure the agar side of the plate is up.</p>
 +
          </li>
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        </ol>
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      </blockquote>
 +
    </blockquote>
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    </p>
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</blockquote>

Latest revision as of 12:11, 20 June 2014

 

PROTOCOLS

6.TRANSFORMATION

  2. 1.Sterilized the enviroment.

  3. 2.Take the compatent cells from -80 ºC thawing on ice.

  4. 3.Add 50 µL of thawed competent cells into 1.5ml tube.

  5. 4.Add 1 - 2 µL of the resuspended DNA to the 2ml tube. Pipet up and down a few times, gently.

  6. 5.Centrifuge tubes at 3000 rpm for 30-45 seconds.

  7. 6.Incubate the cells on ice for 45 minutes.

  8. 7.Heat the tubes in the 42 C heat block for a 80 seconds.

  1. 8.Incubate the cells on ice for 5 minutes.

  2. 9.Add 450 μl of LB broth (make sure that the broth does not contain antibiotics and is not contaminated) to each transformation

  1. 10.Incubate the cells at 37ºC for 1 hour while the tubes are shaking.

    11.Label petri dish with LB agar and the appropriate antibiotic.

  2. 12.Plate 135 µl of the transformation onto the dishes, and spread.

  3. 13.Incubate the plates at 37ºC for 12-14 hours, making sure the agar side of the plate is up.

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