Team:Calgary/7 July 2010


Wednesday July 7, 2010

Wells 2-9 Jeremy's I0500+B0034 with and without restriction enzymes. Wells 11-19 Dev's E0040+B0015 with and without enzymes.


  • Today I finalized the enzymes that we are going to be using for the multiple cloning sites in our transcription/ translation testing circuit. I also had an ethics meeting with Dev and Raida. We talked about what previous iGEM teams have done in terms of Ethics as well as what we are going to be presenting at Suffield next week. I strated reading a few papers on snythetic biology ethics to come up with soe quetsions for next week. Today I also made ovenright cultures of Himika's E1010-B0015 construct as well as her E1010 plasmid switch.


  • Today I completed the transformation of my construct (R0040-I3504) and plated them in 6 different Ampicilin Resistant plates.

Procedure: Transformation

  1. Thaw Competent Cells
  2. Add 20 uL DNA
  3. Ice for 1/2 hour, Heat Shock 5 minutes at 37 degrees Celcius, Ice 5 min.
  4. Recover with 250 uL SOC for 30 minutes
  5. Spin down at 14 RPM, Concentrate to approximately 100 uL
  6. Plate 25 uL on plate 1,2,3 and Plate 50 uL on plate 4,5,6
  7. Leave in Incubator overnight for 20 hours

The plates are labeled as following:

  • P1: R0040-I13504 (construct tube C 6 July) 25 uL
  • P2: R0040-I13504 (construct tube D 6 July) 25 uL
  • P3: R0040-I13504 (construct tube F 6 July) 25 uL
  • P4: R0040-I13504 (construct tube C 6 July) 50 uL
  • P5: R0040-I13504 (construct tube D 6 July) 50 uL
  • P6: R0040-I13504 (construct tube F 6 July) 50 uL

Then I left the plates in the Incubator which is at 37 degrees Celcius. I will leave it there until 10:00 am tomorrow (July 8th), which would let the cells grow for 20 hours.


Today I completed the plasmid switch for I0500 into pSB1AK3, mini prep I0500+B0034 with Sigma-Aldrich and finished a restriction digest of EcoRI and PstI on I0500+B0034

  • 37 degrees for 1 hour and heat kill 80 degrees for 20 minutes
  • ran on 1% agarose gel at 90V


Today, I drafted the potential budget for this year's project. The total expenses this year are projected to amount to $64 500 with that money being used towards paying students, travelling and lodging at MIT, paying for laboratory materials, and buying T-shirts as well as a mascot suit. Our current financial status is sufficient to cover approximately 80% of this with $54 600 in total funds. These have been provided by BioAlberta, Alberta Innovates Technology Funds, O'Brien Student Centre, and Alberta Heritage Foundation for Medical Research.

As well, in the morning, Dev and I used the Sigma Aldrich GenElute Plasmid Preparation kit and modified instructions to isolate the plasmids of the constructions of I0500 - B0034 and E0040-B0015 as well as the parts of I0500, I13507, and E1010.

In the afternoon, I contacted a General Manager from New England Biolabs about possible sponsorship and received a positive response. They will send some enzymes to us in the coming week.


The overnight colonies of the cpxp in pSB1AK3 was pink, which means it was expressing RFP and the construction was unsuccessful. We have decided it would be best to wait for the new primer to come so that we can create new PCR product and construct with that because it contains the entire biobrick suffix, which eliminates the risk of scarring. Today I prepared slides for a presentation to our supervisors on Friday, also I attempted to get into contact with my local newspaper in Bow Island to see if they will do a story on iGEM, no response yet but I’m confident we can get in.


The overnight colonies were pink, save for a single colony. This suggests that RFP was produced, meaning the plasmid switch was unsuccessful. Upon checking the original plates, even the colony that did not turn pink in the culture was pink. Baffling, no doubt, but the conclusion is clear: continuing to use XbaI and SpeI as plasmid insertion sites will definitely be much more trouble than it is worth. Therefore, we ordered in a new reverse primer that also contained the NotI and PstI sites. This will hopefully make the plasmid switch directional.