Imperial College/14 August 2008

=14 August 2008=

Cloning

 * Parts taken from the registry and transformed by electroporation into XL1-Blue E.coli along with XL1-Blue controls and grown on Kanamycin and Ampicillin plates. Parts taken:
 * C0012 (LacI)
 * J31005 (Chloraphemicol acetyltransferase)
 * B0015 (Double terminator)
 * J04630 (GFP and Double Terminator)
 * I13401 (mRFP and Double Terminator)

B.subtilis

 * The transformation protocol 2 was carried out today, | Click link here for protocol
 * The basic principle of this protocol is that competent cells are prepared by growing a culture to a high O.D.600 and then performing electroporation on these competent cells.
 * Today we grew the competent cells, we used an O.D.600 of 1.5 before we harvested them. In addition electroporation was carried out using the plasmid pDR110. Previously we had mini-preped this to give a stock of 40ng/ul. We transformed with 40ng, 120ng, 200ng and 400ng in a volume of 10ul (water was used to make up to 10ul).
 * Transformed cells were plated out and placed into the 30oC incubator.

Dry Lab

 * Completed A More Complex Example of Bayesian Parameter Estimation of Tutorial 2.
 * Sourced for better tracking algorithm, found SpotTracker which is more accurate than ParticleTracker.
 * Went for microscope training, obtained 2 x videos on B.Subtilis motility, stored on server with James.
 * Discussion with Dr. Suhail ( from Structural Bioinformatics Group,Imperial College London) on the computing requirements to analyse the cells motility. We will be allocated a linux 64-bit workstation, that we will be able to access remotely via SSH.

Microscope

 * Chris, Clinton, James and Prudence received microscope training on Widefield 1, recording two 60s videos of B.subtilis swimming for analysis
 * Determined that best results would be obtained by bringing a B.subtilis overnight culture to the microscope facility and diluting 100 fold.