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Team:Nevada/Week 16
From 2012.igem.org
Protocols | Week 1 | Week 2 | Week 3 | Week 4 | Week 5 | Week 6 | Week 7 | Week 8 | Week 9 | Week 10 | Week 11 | Week 12 | Week 13 | Week 14 | Week 15 | Week 16 | Week 17 | Week 18 | Week 19 |
Contents |
September 4
- Joe:
- gel 186 shows colonies 4, 6, 7, 8 are good
September 5
- Jeremiah & Chris:
- Colony PCR check 12 colonies
- Check on a gel (gel #187)
- Samples 8 and 9 cultured in LB + Amp
September 6
- Joe:
- miniprep of colonies 7, 8 RFP-SBP
- glycerol stock of SBP-RFP in Top10 cells
- digestion of minipreps by EcoRI and PstI
- Transform to BL21 cells
- culture more RFP-SBP in Top10 cells
- Jeremiah & Chris:
- Miniprep plasmid
- Colony check plasmid again to make sure it’s not a false positive
- Check on gel (gel #188)
- Transform this plasmid (L. Arabanose promoter with TBP+ insert) into BL21 cells
- Used Amp antibiotic on plates
September 7
- Joe:
- Culture colony of RFP-SBP in BL21 cells
- heat inactivate RFP-SBP(EcoRI/PstI) - gel 189 confirms good
- Michelle:
- Culture M2-BL21 colonies from LB AMP-CM to make glycerol stocks. M2-BL21 along with purified desalted SBP-LRP :::protein used to carry out Western Transfer Blot and SDS-PAGE Coomassie staining procedure. Lysis of M2-BL21 :::pellet with protein inhibitor mixed with PBS and B-Mercaptoethanol 48.7%.
- Justin and Dafne:
- Large scale expression of SBP-B12 protein in BL21 cells
