Latest revision as of 08:51, 10 September 2009

Project progress

A miniprep and RD (with EcoRI and PstI) were performed on the four colonies that were ented on Sunday.
- Nanoprop results
- RD results on gel: the right signals were detected

LigA was plated on LB medium. This will be the "motherplate" from which the glycerol stock and the testing cultures will be taken from.
Electroporation of in competent cells. It was plated and put overnight in the incubator.
In order to be able to grow vector , which contains the toxic ccdB gene, we need special cells which carry the gyrA462 mutation. This is strain DB3.1 from E.coli. These were plated and put in the incubator overnight.

Electroporation of SAMS I, SAMS II AND EF II
Restriction digest of ech and fcs from Saturday was tested and showed 6 signals with varying lengths for ech and 1 very large signal for fcs, both inconsistent. To test for errors in the restriction sites we cut each vector with 1 restriction digest for each of the sites. Giving a total of 4 tests for ech and 4 for fcs.
- Restriction at 37°C overnight.
For ligation of SAMS III RD (from Thursday) and TER II we need 44,4 µl of SAMS III which we don't have thus we cut two additional volumes with SAMS I overnight
- SAMS I A RD and SAMS I B RD

TOPO cloning showed only blue colonies so we searched for alternatives. We did resaerchwork if it was maybe possible with a puc-vector
The mutation of rpoA was again checked but gelelectrophoresis showed no result
X,Y and K (the igemvector) were cut with EcoR1 and Pst1.
new ligation was done with the first cuttingproduct so today it was electroporated and plated.
PCR was done for W to get more basicproduct to work with

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