Construction

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<font size=4>< [[Aug11-Sep7]] | [[Sep8-14]] ></font>
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<font size=4>< [[Aug11-Sep7]] | [[Sep8-14]] | [[Sep15-26]]></font>
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     <li>Learn to use new DH5a (with no LacI expression) for testing
     <li>Learn to use new DH5a (with no LacI expression) for testing
     <li>Post up new protocols
     <li>Post up new protocols
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</ul>
 
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[http://2006.igem.org/University_of_Toronto_2006 Home]
 
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== September 26, 2006 ==
 
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'''Charles:'''
 
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<ul>
 
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    <li>Prepared o/n of UT2 (2 vials) and UT3 (2 vials) for a freezer stock
 
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</ul>
 
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'''Andy:'''
 
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<ul>
 
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    <li>Prepared o/n of UT3 BCDE for mini-preps
 
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    <li>transformed and plated P0452 (2005) and P0456 (2005)
 
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</ul>
 
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'''To-Do List:'''
 
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<ul>
 
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    <li>Make freezer stock and minipreps of above tubes and check lengths along with P0352 (2005) and P0152 (2005)
 
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    <li>Make more plates, autoclave tips, make more LB
 
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    <li>Post up new protocols, include part length and resistance info in spreadsheet
 
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</ul>
 
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[http://2006.igem.org/University_of_Toronto_2006 Home]
 
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== September 22, 2006 ==
 
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'''To-Do List:'''
 
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<ul>
 
-
    <li>Make frozen stock of UT2 and UT3
 
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    <li>Prepare o/n for UT3 BCDE for mini-prep
 
-
    <li>Post up new protocols, include part length and resistance info in spreadsheet
 
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    <li>Make more plates and re-plate P0452 (2005) – AK, and P0456 (2005) – AC
 
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</ul>
 
-
 
-
[http://2006.igem.org/University_of_Toronto_2006 Home]
 
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== September 21, 2006 ==
 
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'''Charles:'''
 
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<ul>
 
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    <li>Made 22 competent cells
 
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    <li>Tested Module 1 using varying concentrations of IPTG (0, 2, 20, 200, 2000 uM) to induce GFP
 
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        (E0240 (2005)) and mRFP (I13507 (2005))
 
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    <table>
 
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        <tr><td colspan=2>[[Image:Table_GFP.jpg]]</td>
 
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        <tr><td>[[Image:Graph1_GFP.jpg]]</td><td>[[Image:Graph2_GFP.jpg]]</td>
 
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        <tr><td>[[Image:Table_mRFP.jpg]]</td><td>[[Image:Graph_mRFP.jpg]]</td>
 
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    </table>
 
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    <li>Data indicates that the GFP and mRFP as well as the LacI+ pL promoter (R0011) are functional
 
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        (Does not indicate functionality of the pBad/AraC promoter and the inverter)
 
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    <li>Some of the replicates seem to be outliers and if excluded, the regression increases
 
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        significantly
 
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    <li>DH5a-z1 are known to fluoresce, so the fact that untransformed cells glow brighter is not a
 
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        big concern
 
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    <li>Next step is to transform UT2 and UT3 into LacI negative cells then:
 
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        <ul>
 
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            <li>Characterize the fluorescence under varying Arabinose concentrations.
 
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            <li>Vary the temperature to verify the temperature sensitivity of LacI ts and its affects
 
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                on reporter expression
 
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        </ul>
 
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</ul>
 
-
 
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'''To-Do List:'''
 
-
<ul>
 
-
    <li>Post up new protocols, include part length and resistance info in spreadsheet
 
-
    <li>Make more plates and re-plate P0452 (2005) – AK, and P0456 (2005) – AC
 
-
</ul>
 
-
 
-
[http://2006.igem.org/University_of_Toronto_2006 Home]
 
-
 
-
== September 20, 2006 ==
 
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'''Natalie, Andy:'''
 
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<ul>
 
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    <li>Prepared o/n growth of  UT3 BCDE for mini-prep, DH5a for competent cells, and UT2 – 3,4,5, UT3 – 7,8,9 and DH5a x 2 (control) for making fluorescence measurements.
 
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    <li>Checked lengths of miniprep UT2 ABCDEF, UT3 A (possibly unreliable due to 2 day o/n growth), P0352 (2005), and P0152 (2005).
 
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</ul>
 
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'''To-Do List:'''
 
-
<ul>
 
-
    <li>Make 24 competent cells
 
-
    <li>Miniprep UT3 BCDE and test lengths
 
-
    <li>Make more plates and re-plate P0452 (2005) – AK, and P0456 (2005) – AC
 
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    <li>Do measurements on UT2 and UT3
 
-
    <li>Post up new protocols, include part length and resistance info in spreadsheet
 
-
</ul>
 
-
 
-
[http://2006.igem.org/University_of_Toronto_2006 Home]
 
-
 
-
== September 18, 2006 ==
 
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'''Charles:'''
 
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<ul>
 
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    <li>Only mini-prepped UT3 A because they were grown for two o/n  (Just want to see what happens)
 
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    <li>Prepared new o/n for UT3 BCDE as well as 1 DH5a-z1, 2 UT2 and 2 UT3 for testing tomorrow. And
 
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        1 DH5a-z1 for making competent cells
 
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</ul>
 
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'''To-Do List:'''
 
-
<ul>
 
-
    <li>Make 24 competent cells
 
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    <li>Test the lengths of UT2 ABCEDF, P0352 (2005), P0152 (2005), and UT3 BCDE
 
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    <li>Make more plates and re-plate P0452 (2005) – AK, and P0456 (2005) – AC
 
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    <li>Prepare new o/n for UT3 BCDE – for mini-prep
 
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    <li>Prepare o/n in the following quantities: 3 DH5a-z1, 3 UT2 and 3 UT3
 
-
</ul>
 
-
 
-
[http://2006.igem.org/University_of_Toronto_2006 Home]
 
-
 
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== September 16, 2006 ==
 
-
 
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'''Charles:'''
 
-
<ul>
 
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    <li>Made 4 replications of UT2 in 200 uM, 100 uM, 50 uM, 25 uM and 0 uM of IPTG.
 
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    <br><br><center>[[Image:Module01_Test01_Raw.jpg]]<br>
 
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        [[Image:Module01_Test01_Graph.jpg]]</center><br>
 
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        <ul>
 
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            <li>OD of cells is too high, need to be <= 1.2
 
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            <li>Need a control of untransformed DH5a-z1 to compare the relative fluorescence
 
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        </ul>
 
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    <li>Made frozen stock (2 vials each): UT01, I13507 (2005), B0015 (2005), B0031 (2005)
 
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    <li>Mini-prepped UT2 ABCEDF, P0352 (2005) and P0152 (2005)
 
-
</ul>
 
-
 
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'''To-Do List:'''
 
-
<ul>
 
-
    <li>Test the lengths of UT2 ABCEDF, P0352 (2005) and P0152 (2005)
 
-
        <ul>
 
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            <li>Prepare an o/n growth of the “best” UT2 for testing
 
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            <li>Prepare an o/n growth of regular DH5a-z1
 
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        </ul>
 
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    <li>Mini-prep UT03 ABCD and test their lengths along with P00352 (2005) and P0152 (2005)
 
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    <li>Re-plate P0452 (2005) - AK, and P0456 (2005) - AC
 
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    <li>Make 24 competent cells
 
-
</ul>
 
-
 
-
[http://2006.igem.org/University_of_Toronto_2006 Home]
 
-
 
-
== September 15, 2006 ==
 
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'''Andy, Charles, Natalie, Stan, Elliott:'''
 
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<ul>
 
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    <li>Checked and transformed I+J+I plasmids (Bands: ~6000-5000, 3500-2500 (smear))
 
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    <li>Made competent cells (although, may have to repeat)
 
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    <li>Prepared o/n growths of I+J+E (12 vials), P0152 (2005) and P0352 (2005). P0456 (2005) and P0452 (2005) did not seem to grow
 
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        very well so left them in the incubator for a 2nd night
 
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    <li>Made 8 Amp plates
 
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</ul>
 
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'''To-Do List:'''
 
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<ul>
 
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    <li>Mini-prep P0152 (2005), P0352 (2005) and 6 of the I+J+E
 
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    <li>Try to get J04450 (2006) working again.
 
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    <li>Check lengths of R0011 (2005/6), if no good, then obtain R0010 (2005/6) from Registry
 
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    <li>Make freezer stock (2 each) of I+J, B0015 (2005), B0031 (2005), and I13507 (2005) and put in -80C freezer stock
 
-
    <li>Test the fluorescence of I+J+E
 
</ul>
</ul>
[http://2006.igem.org/University_of_Toronto_2006 Home]
[http://2006.igem.org/University_of_Toronto_2006 Home]

Revision as of 16:44, 30 September 2006

< Aug11-Sep7 | Sep8-14 | Sep15-26>

September 29, 2006

Charles:

  • Prepared o/n of competent cells (DH5a-z1 and DH5a), and UT3 ABCD to miniprep again

To-Do List:

  • Make new competent DH5a-z1
  • Prepare o/n of regular DH5a
  • Plated P0 (2006) using new competent cells
  • Double digest UT1 (host) with SpeI/PstI in Buffer 2 and double digest P0352/P0456 (2005) with XbaI/PstI in Buffer 3

[http://2006.igem.org/University_of_Toronto_2006 Home]

September 28, 2006

Andy:

  • Miniprep P0456 (2005)
  • Checked the lengths (EcoRI and PstI, Buffer EcoRI): UT3 ABCDE, P0456 (2005) and P0352 (2005)
  • Transformed P0152, P0352, P0452, and P0456 (2006). [Plated on Amp plates, because no more Amp/Kan plate]

To-Do List:

  • Plate DH5a cells
  • Make LB Broth, plates and print protocols
  • Digest P0352 and P0456 (2005) and ligate with UT1
  • Prepare o/n of UT3 ABCDE again and try to miniprep again

[http://2006.igem.org/University_of_Toronto_2006 Home]

September 27, 2006

Charles:

  • Made freezer stocks of UT2 x 2 and UT3 x 2

Andy, Natalie:

  • Mini-preppred UT3 B(AB) C(BB) D(BC) E(CC)
  • Transformed and plated P0452 (2005) (On Amp rather than proper AK)
  • Checked lengths of UT3 BCDE (inconclusive due to anomalous bands, D appears promising - yellow rating), P0152 (2nd check, Extra band, wrong insert length), P0352 (2nd check, not the best bands, correct lengths - changed rating from red to yellow)
  • Prepared o/n of P0456 (2005) for Miniprep
  • Added part lengths and resistance info in spreadsheet

To-Do List:

  • Miniprep P0456 (2005)
  • Take P0456, P0452, P0152, P0352 from 2006 registry
  • Check lengths of UT3 BCDE and P0352 again
  • Make more plates, make more LB
  • Learn to use new DH5a (with no LacI expression) for testing
  • Post up new protocols

[http://2006.igem.org/University_of_Toronto_2006 Home]

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