Week of 6/14

From 2010.igem.org

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*We then cut out each available band, purified, ligated, and finally transformed.
*We then cut out each available band, purified, ligated, and finally transformed.
 +
 +
==June 15th==
 +
 +
 +
*Flasks mini prepped
 +
    -J37033 (LuxR + RBS)
 +
    -C0261 (LuxI + RBS)
 +
 +
*All of the mini preps were digested with the following enzymes:
 +
    -J37033 (XP)
 +
    -C0261 (XP)
 +
 +
*The digests were put into a 37 degree Celsius water bath for an hour. The reaction was heat killed for 20 minutes in an 80 degree Celsius water bath.
 +
 +
*The two parts were then run on a gel in this order: Ladder, PFTV (another student's project), C0261, and J37033.
 +
 
<center>[[Image:6-15-10.jpg]]</center>
<center>[[Image:6-15-10.jpg]]</center>
 +
 +
 +
 +
*This time the parts came out very vibrantly. We extracted C0261, did the gel purification, ligated, and transformed.
 +
 +
==June 16th==
 +
 +
 +
*Flasks mini prepped
 +
    -R0010 (pLac)
 +
    -C0062 (LuxR)
 +
 +
*All of the mini preps were digested with the following enzymes:
 +
    -R0010 (XP)
 +
    -C0062 (XP)
 +
 +
*The digests were put into a 37 degree Celsius water bath for an hour. The reaction was then heat killed for 20 minutes in an 80 degree Celsius water bath.
 +
 +
*After the heat kill, an Agarose gel was run. The parts run on the gel were Ladder, R0010, and C0062.
 +
 +
<center>[[Image:6-16-10.jpg]]</center>
<center>[[Image:6-16-10.jpg]]</center>
 +
 +
 +
 +
*After the gel was finished running, we extracted the necessary bands and purified them.
 +
 +
*We made notes of possible ligations:
 +
    -Lux Promoters <-> C0261 (LuxI)
 +
    -Lux Promoters <-> K274100 (Pigment)
 +
    -Constitutive Promoters <-> J37033 (Lux R)
 +
    -CI Promoters (K091107) <-> Lysis (K112022, K112808)
 +
 +
*Goal for June 17th:
 +
    -Ligate B0034 <-> C0051
 +
                      C0012
 +
                      C0062
 +
 +
==June 17th==
 +
 +
 +
*Flasks mini prepped:
 +
    -J24679
 +
    -K091107
 +
    -R0061
 +
    -J37033
 +
    -K081007
 +
    -LacI + RBS
 +
 +
*All of the mini preps were digested with the following enzymes:
 +
    -K081007 (XP) (Phosphatase)
 +
    -K091107 (SP)
 +
    -J37033 (XP) (Phosphatase)
 +
    -R0061 (SP)
 +
    -J37033 (PCR) (XP)
 +
    -C0261 (PCR) (XP)
 +
 +
*The digests were put into a 37 degree Celsius water bath for one hour. The reaction was then heat killed for 20 minutes in an 80 degree Celsius water bath.
 +
 +
*After the heat kill, an agarose gel was run in the order of Ladder, J37033 (PCR), K274100, C0261 (PCR), R0061, K091107, and then P, T, V (another project).
<center>[[Image:6-17-10.jpg]]</center>
<center>[[Image:6-17-10.jpg]]</center>
 +
 +
 +
 +
*Once the gel was finished we extracted and purified the parts.
 +
 +
*We made competent cells, SOB media, and Tet plates.

Latest revision as of 15:40, 5 August 2010

Contents

June 14th

  • Flasks mini prepped 
   -J37033 (LuxR + RBS)
   -C0261 (LuxI + RBS)
   -R0061 (Lux Promoter)
   -J23116 (Constitutive Promoter)
   -R0063 (Lux Promoter)
   -K145150 (Lux Promoter)
   -K091146 (Lux Promoter)
   -K274100 (Pigment)
   -J23108 (Constitutive Promoter)
  • All of the mini preps were digested with the following: 
   -J37033 (XP)
   -C0261 (XP)
   -R0061 (ES)
   -J23116 (ES)
   -R0063 (ES)
   -K145150 (ES)
   -K091146 (ES)
   -K274100 (ES)
   -J23108 (ES)
  • We ran the digest as usual: 1 hour in the water bath, 20 minutes heat kill. We did not phosphotase any of these as you can see all of these parts appear on the gel.
  • The order was Ladder, J23108, K274100, K091146, K145150, R0063, J23116, R0061, C0261, and J37033.



6-14-10.jpg


  • This gel came out well with the exception of K145150.
  • We then cut out each available band, purified, ligated, and finally transformed.

June 15th

  • Flasks mini prepped 
   -J37033 (LuxR + RBS)
   -C0261 (LuxI + RBS)
  • All of the mini preps were digested with the following enzymes: 
   -J37033 (XP)
   -C0261 (XP)
  • The digests were put into a 37 degree Celsius water bath for an hour. The reaction was heat killed for 20 minutes in an 80 degree Celsius water bath.
  • The two parts were then run on a gel in this order: Ladder, PFTV (another student's project), C0261, and J37033.


6-15-10.jpg


  • This time the parts came out very vibrantly. We extracted C0261, did the gel purification, ligated, and transformed.

June 16th

  • Flasks mini prepped 
   -R0010 (pLac)
   -C0062 (LuxR)
  • All of the mini preps were digested with the following enzymes: 
   -R0010 (XP)
   -C0062 (XP)
  • The digests were put into a 37 degree Celsius water bath for an hour. The reaction was then heat killed for 20 minutes in an 80 degree Celsius water bath.
  • After the heat kill, an Agarose gel was run. The parts run on the gel were Ladder, R0010, and C0062.



6-16-10.jpg


  • After the gel was finished running, we extracted the necessary bands and purified them.
  • We made notes of possible ligations: 
   -Lux Promoters <-> C0261 (LuxI)
   -Lux Promoters <-> K274100 (Pigment)
   -Constitutive Promoters <-> J37033 (Lux R)
   -CI Promoters (K091107) <-> Lysis (K112022, K112808)
  • Goal for June 17th: 
   -Ligate B0034 <-> C0051
                     C0012
                     C0062

June 17th

  • Flasks mini prepped: 
   -J24679
   -K091107
   -R0061
   -J37033
   -K081007
   -LacI + RBS
  • All of the mini preps were digested with the following enzymes: 
   -K081007 (XP) (Phosphatase)
   -K091107 (SP) 
   -J37033 (XP) (Phosphatase)
   -R0061 (SP)
   -J37033 (PCR) (XP)
   -C0261 (PCR) (XP)
  • The digests were put into a 37 degree Celsius water bath for one hour. The reaction was then heat killed for 20 minutes in an 80 degree Celsius water bath.
  • After the heat kill, an agarose gel was run in the order of Ladder, J37033 (PCR), K274100, C0261 (PCR), R0061, K091107, and then P, T, V (another project).


6-17-10.jpg


  • Once the gel was finished we extracted and purified the parts.
  • We made competent cells, SOB media, and Tet plates.




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