Team:Caltech/Week 4

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 [http://partsregistry.org/Part:BBa_V1012 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 [http://partsregistry.org/Part:BBa_V1012 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 [http://partsregistry.org/Part:BBa_V1012 V1012] in case we need to retry the competence procedure.
• Tested pigment production of cells containing [http://partsregistry.org/Part:BBa_K274100 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.

Thursday 7/8

• The [http://partsregistry.org/Part:BBa_V1012 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 [http://partsregistry.org/Part:BBa_M30109 M30109] for assembly tomorrow, along with cultures of [http://partsregistry.org/Part:BBa_J04450 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.

Friday 7/9

Weekend 7/10-11