Imperial College/31 August 2008

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31 August 2008

Wet Lab

Cloning

  • Whenever the sequences from GeneArt arrive, priority becomes the rapid cloning of sequences into Biobricks to begin construction
Monday
  • Test all primers for PCR cloning from pDR111 using taq to ascertain optimal consitions.
  • If time allows, test genomic primers using B.subtilis chromosome sample 1 or using single colony PCR
Tuesday
  • Carry out PCR cloning with Pfu, purify and clone DNA sequences into Biobricks
Wednesday
  • Test orientation of inserts to find a correct Biobrick containing host to culture overnight for midiprepping
Thursday & Friday
  • Hopefully, construction begins...
B.subtilis
Monday
  • Prepare all media and plates needed for the rest of the week,
  • Carry out single colony PCR on B.subtilis transformed previously with pDR111 using verification primers. These verification primers work on the basis of flanking the chromosomal integration sites allowing the integration sites and any inserted elements to be amplified.
Tuesday
  • Prepare overnight culture for the B.subtilis growth curve
  • Perform a transformation using linear DNA derived from pDR110 or pDR111
Wednesday
  • 2nd trial of the B.subtilis growth curve and OD600 vs c.f.u. experiment
  • Perform single colony PCR with the B.subtilis transformed yesterday using verifications primers.
  • Begin to try to optimise the protocol for B.subtilis transformation protocol with linear DNA. This includes the volume of transformed cells, amount of DNA, the density of competent cells and length of DNA incubation.
Thursday


Friday
  • Carry on to optimise the protocol for B.subtilis transformation protocol with linear DNA

Dry Lab

ODE team Weekly plan
  • Develop the MATLAB simulation of the genetic circuitry: simulate behaviour of the constituive and inducible gene expression of the genetic parts.
  • Develop the characterisation of the B.Subtilis chassis; simulate an ODE system regarding the growth of the B.Subtilis (model the process of the diffusion of nutrients into the bacteria, additionally simulating the lag phase and the stationary phase of the growth)
  • Present a list of parameters for wet lab



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