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Team:Brown/Notebook/August12
From 2010.igem.org
Thursday, August 12, 2010
Miniprep of Lovtwo
Used white culture from XL1B blue-white selection. Final concentration was 227.1ng/ul.
Digest of Lovtwo miniprep with EcoRI
50ul total volume:
- 5ul NEB buffer 2
- 2ul DNA
- 0.5ul 100x BSA
- 1ul EcoRI
- 41.5ul dH2O
Vortexed, spun down, incubated @37C at 12:18PM
Gel of AraC, CI, Mnt
Gel of AraC, Mnt, CI, LovTwo
16ul DNA, 4ul buffer per well. AraC little tube only had 10ul DNA (labeled).
Glycerol Stocks
Made glycerol stock of BL-21 line that contains J04421 LacI reporter
AraC RFC25 gel extract
Gel extract of AraC (RFC25) PCR product resulted in a low yield of only 3.4ng/ul. Will re-PCR
Re-PCR of AraC with RFC25 sites
20ul total:
- 10ul 2x Master mix
- 1ul fwd primer
- 1ul rev primer
- 1ul DNA
- 7ul dH2O
Overnights
Picked culture from plate with expression vector J13002. Grew overnight in LB Amp+ liquid culture
