Team:Arizona State/Notebook


May: Week 1

  • First ASU iGEM 2013 with new team members
  • Introduction to the competition, registration logistics, summer schedules
  • Idea Brainstorming
  • Review of biobrick cloning, Golden Gate assembly, safety training

Week 2

  • Idea Brainstorming--narrowed down to two project ideas

Safety Training:

  • Biosafety and Bloodborne Pathogens
  • Lab Safety
  • Fire Safety
  • Recombinant DNA Safety
  • Hazardous Waste Management

June: Week 3

  • Started making amp plates in preparation for experimentation

Week 4

  • Started growing up colonies of E. coli K12 MG1655, BL21 (DE3),and N10 B strains along with glycerol stocks
  • Performed a restriction, ligation and transformation of the three plasmids and grew them up on a plate
  • No colony growth of any of the plasmids

Week 5

  • Performed a PCR amplification of the cadA promoter along with a gel screen that found the band at the appropriate band length
  • Transformed and plated both J61100 and E1010 with no growth on either plates
  • Gel screen of yebF without the stop codon never showed up on the gel. Ran transformation again.
  • Restricted and ligated cadA and BN promoters

Week 6

  • The gel screen of yebF without the stop codon was finally successful
  • Transformation of INPNC-MCS and RFP control onto N10B cells was successful
  • No growth from transformation of E0020, E0030, and E0040
  • Created liquid cultures of E0240 and pCadA in LB Chloro.

July: Week 7

  • Performed restriction digest of pCadA, pBN, and YebF
  • Performed seven different ligations
  • Miniprepped and nanodropped E0240 and pCadA.
  • Miniprepped YebF+GMCSF and PelB leader sequence.
  • Flow cytometry indicated that there was no GFP
  • Transformed J23100 yet no plate growth

Week 8

  • Ordered Antigen primers
  • Ligation and transformation of PelB+GMCSF+pSB1C3/4K5
  • Miniprep showed no successful ligation and transformation

Week 9

  • PCR of MelanA and Flu M1 worked via confirmation from gel screen
  • Nissle would not grow
  • Transformed I13522 into N10B on LB Amp

August: Week 11

  • Grew up six cultures of ligations
  • Miniprepped the PSB1c3 backbone and grew up additional backbone in cultures
  • Transformed the GFP and GMCSF in the backbone
  • Ran LLO pcr on gel and all had correct band lengths

September: Week 15

  • Made ligations of FLU+1c3, MelA+1c3, Flu+J04..., MelA+J04...
  • Made liquid cultures of non-red colonies from FLU+1c3, MelA+1c3, Flu+J04..., MelA+J04 transformation
  • Biobricks made and sent: LLO with promoter RBS, YEb-GMCSF, GMCSF, FLU1, and MelanA