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Week9 8/8/10-8/14/10
From 2010.igem.org
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Miniprepped circular backbone plasmid | Miniprepped circular backbone plasmid | ||
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===8/11/10=== | ===8/11/10=== | ||
Revision as of 18:25, 16 October 2010
Contents |
8/9/10
Bought Chitosan powder to use as a positive control for calcofluor white --> glowed bright blue/green when stained
Stained (pMal-CHS3) ligation products for the presence of chitin --> ligation 1/tube 3 had slight fluorescence
Used UV-vis Spectroscopy to measure chitin concentrations in ________
Grew bakers yeast to test for presence of chitin
Colony PCR on (pMal-CHS3)
CP-LacPI part testing
8/10/10
Ran gel for colony PCR no positives
Ran PCR for CHS3 insert into pMal vector
Stained varying concentrations of chitosan (.25g/ml, 25ug/ml, 5ug/ml, 1ug/ml) and (pMal-CHS3) cells --> cells showed no signs of chitin
Digested __________ (sean)
Ligated (CP-LacPi)-(GFP) in Tet backbone
Made LB minimum media for electroporated cells
Miniprepped circular backbone plasmid
8/11/10
Digested and ran a gel of CHS3 and backbone plasmids (C,T,K,A)
Tested restriction enzymes by digesting circular plasmids with each enzyme individually --> should see 1 2500kb band
Organized the DNA samples in our -20.
8/12/10
Weekly meeting
- Went over our gels --> identified faulty parts (holin1, holin2, CHS3-pMal, CP-LacPI)
- Plan to re-kit to stock and reassemble
Decided to use an alternative stain because calcofluor results in too much background and our confocal microscope can not excite in the UV spectrum
8/13/10
Methanol fixation of yeast and E.Coli cells --> stained with rhodamine-conjugated chitin probe (allowed to incubate overnight)
Competency of ChiA knockouts = 2.5x10^7 and 5.0x10^6
Cycles 1 of File:MAGE for tqsA Knockouts.
