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Team:Caltech/Week 4 - 2010.igem.org
 

Team:Caltech/Week 4

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* Completed schematics for our printing process, including a detailed gene network diagram. We will begin assembly of the 3D genetic constructs tomorrow.
* Completed schematics for our printing process, including a detailed gene network diagram. We will begin assembly of the 3D genetic constructs tomorrow.
* Results of the pigment test with [http://partsregistry.org/Part:BBa_K274100 K274100]: None of the cells seemed to have created the red pigment we were looking for.
* Results of the pigment test with [http://partsregistry.org/Part:BBa_K274100 K274100]: None of the cells seemed to have created the red pigment we were looking for.
 +
** It seems likely that that all the cells died.
 +
** Did not have a visible pellet of cells in the glycerol and agar solutions after centrifugation.  The solutions were probably too viscous. 
==Friday 7/9==
==Friday 7/9==

Revision as of 19:59, 9 July 2010


iGEM 2010



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Contents

Monday 7/5

Note: Today was another Institute Holiday.

Tuesday 7/6

  • Hydrolyzed linseed oil. The reaction solution will be characterized on Wednesday.

Wednesday 7/7

  • Attempted to make V1012 (strain CP919, KanR, envZ deficient) electrocompetent:
    • Centrifuged overnight 3mL LB-Kan culture at 4C, 16000rcf for 10min.
    • Discarded supernatant. Resuspended pellet in 1.5mL ice-cold water. Centrifuged again.
    • Repeated with 0.75mL ice-cold water.
    • Resuspended pellet in 1.5mL ice-cold 10% glycerol.
    • Flash-froze 100uL aliquots in dry ice/ethanol.
  • Transformed light/lysis ligation product from last week into V1012 via electroporation. (Voltage: 2.5V; time constant: 4.5)
    • Plated 100uL on LB-Tet agar plate & inoculated a 5mL LB-Tet overnight culture.
  • Inoculated 5mL LB-Kan culture of purely V1012 in case we need to retry the competence procedure.
  • Tested pigment production of cells containing K274100 in various solutions.
    • Tested solutions of LB, LB without yeast extract added, 80% glycerol, 100% glycerol, and glycerol with different concentrations of agar.
    • Since the brick is under an arabinose-inducible promoter, two versions of each solution were made - one containing arabinose, and one without arabinose.
    • Pigment production was subjectively measured every hour for five hours. It seemed like there wasn't going to be much pigment produced so the solutions were left overnight in the shaker.

Thursday 7/8

  • The V1012 transformation appears to have worked - the plate is covered in 1000+ colonies.
    • Made freezer stock from parallel liquid culture.
  • Miniprepped the DNA and prepared a sample for sequencing.
  • Started LB liquid culture of M30109 for assembly tomorrow, along with cultures of J04450 to perform a quick lysis test (in preparation for testing the lysis construct).
  • Prepared 2YT and SOB media in preparation for making electrocompetent DH5\alpha cells tomorrow. Also prepared all glassware and other supplies.
  • Completed schematics for our printing process, including a detailed gene network diagram. We will begin assembly of the 3D genetic constructs tomorrow.
  • Results of the pigment test with K274100: None of the cells seemed to have created the red pigment we were looking for.
    • It seems likely that that all the cells died.
    • Did not have a visible pellet of cells in the glycerol and agar solutions after centrifugation. The solutions were probably too viscous.

Friday 7/9

Weekend 7/10-11

 
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