Lab Notebook

23 July 2009

Tim Weenink

Details of the following to be provided tomorrow:

grew up *S in LC

looked at gel

Did !A assembly again with double the amount of backbone (and rest from fridge). Did it with both DH5alpha and homemade top10 chemically competent cells, to compare transformation efficiencies. Also the right concentration of K was used

PCR'd 8 colonies from my transformation plates with VF2 and I-SceI Reverse primers to see if there was incorporation of the I-SceI restriction site

isolated plasmid from *S

Sriram

The agarose gel electrophoresis was run for another 1 hour and 10 minutes at 110 V. Then the gel was immersed in TBE buffer stained with 20µl of Safe Green gel stain for 1 hour and the bands were viewed to check whether we have the right biobricks. By this we confirmed that all the amplified biobricks to be used in Delay were extracted well and they are pure.

For preparing glycerol stocks and plasmid DNA extraction the 7 biobrick colonies [R0010, R0040, J23008, J23031, B0034, B0015, K081013] from the LB agar plates and 2 composite biobricks [S03335, S03473] received from iGEM HQ were cultured in 5 ml tubes with 1xAmp. The 7 biobricks were recultured since we anticipate that we may need more DNA for parts like RBS, Double terminator, etc. in more assemblies and also glycerol stocks were not made for these biobricks.

The 2 composite biobricks [S03335, S03473] were also streaked in LB Agar plates with 1xAmp for future use.

Orr

Prepared TBE and LB Agar mediums using the standard protocols.

Looked at the negative feedback Matlab code in an attempt to write a new code that will enable us to change two variables at a time and check a set of values to find the optimal value for each parameter.

Calin

Gel showed that oriT and promoter miniprep didnt go right. Cultured oriT, promoter, GFP gen in 5 mL tubes. Made 2 GFP plates, an RFP plate, and one with both. Plated parts from MIT (BBa_J23015, BBa_J23055). Added composite part with GFP and oriT to sandbox. Acquired some 20% L-arabinose. Looked into the proofreading PCR kit in the lab for possible use in the knockout (Roche Expand High Fidelity PCR System).