"
Team:ETHZ Basel/Biology/Status
From 2010.igem.org
generation of subparts, linkers and vectors
| Protein | primers | PCR product | storage vector | digest | sequencing | ||
|---|---|---|---|---|---|---|---|
| che proteins | CheY-A | yes | purified | yes | worked | verified | |
| CheY-C | yes | purified | yes | worked | verified | ||
| CheB-A | yes | purified | yes | worked | |||
| CheB-C | yes | purified | yes | worked | |||
| CheR-A | yes | purified | yes | in progress | |||
| CheR-C | yes | purified | yes | in progress | |||
| localizers | TrigF-A | yes | purified | yes | in progress | ||
| MreB-C | yes | purified | yes | in progress | |||
| tetR-A | yes | purified | yes | in progress | |||
| tetO7 | yes | purified | yes | worked | |||
| linkers | PIF3-A | yes | purified | yes | in progress | ||
| PIF3-C | yes | purified | yes | in progress | |||
| PhyB-C | yes | purified | yes | in progress | |||
| reporters | cyFP-A | yes | purified | yes | in progress | ||
| cyFP-C | yes | purified | yes | in progress |
Linker oligos are recieved and don't need to be treated further. They can be used directly for the generation of the BioBricks.
storage and acceptor vectors are available.
PIF3 and PhyB were additionally ordered for synthesis (codon optimized). Not yet recieved.
generation of biobricks
We'll talk to Fabian. Apparently he has a nice protocol to generate acceptor vectors and transform E. coli sequentially in a 96 well plate.
testing of biobricks
testing of che-fusions
We're currently trying to establish an assay to test for the chemotactic behavour of E. coli. A first try with a "syringe test" failed. We'll redo the test and are looking for other possibilites. Maybe a swarm test?
- tested biobricks: che-fusions
- negative control: E. coli mutated for the accoding che-protein
- positive control: wild type E. coli
testing of localicer-fusions
We are planning to use a fluorescence microscope in order to visualize the distribution of localizer-cyFP fusions within a single E. coli.
testing of couplers
We're planning to use the setup as for the testing of the localizers.
- tested biobricks: localizer-PhyB and PIF3-cyFP, localizer-PIF3 and PhyB-cyFP
Images should look as in testing of localizer-fusions. Preliminary clarificaiton of the suitability of the approach are to be done.
bringing E. lemming alive
timetable
