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User:University of Washington/27 June 2008
From 2008.igem.org
Contents |
Yeast Shuttle Plasmid
- 1.5 mL 15% glycerol stocks of the overnight BY4741 yeast growth cultures were made by combining 563 uL of 40% glycerol and 937 uL yeast culture, then stored at -80 degrees Celsius.
Conjugation
Mini Prep
·R0062 (LuxR promoter)
Glycerol
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Sequencing
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LuxR from AraC and TetR
- Made glycerol stock
- Did mini Prep
- Submitted DNA for sequencing reaction and analysis (only for AraC)
LuxR from pLac
- I0462 was again transformed into MG strain again, and plated on AMP.
BioBrick Promoter Measurements
- A strain of TOP10 E. coli was obtained and thawed on ice, along with a strain which had been previously transformed (at MIT) with the fully-assembled reference promoter-RBS-GFP gene-backbone plasmid (I20260).
- 40 uL aliquots of the untransformed cells were transferred into five different tubes. A 40 uL aliquot of the already-transformed cells was similarly transferred into its own tube.
- 2 uL each of plasmid solutions I20260, J23150, J23151, and J23102 were added to four separate aliquots of TOP10 cells.
- The TOP10-pDNA mixtures, the pretransformed TOP10 cells, and the remaining negative control with only untransformed TOP10 cells were incubated on ice for 5 minutes, then incubated in a water bath at 42 degrees Celsius for 30 seconds, then transferred back to ice.
- 250 uL of SOC media was added to each of the TOP10 tubes, then incubated at 37 degrees Celsius on a rotator for 1 hour.
- 100 uL of the each of the transformed cells were placed on Kanamycin plates, then left at room temperature over the weekend to allow for slow growth.
Team:University_of_Washington/Notebook#Notebook
