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Team:Hawaii/Notebook/2008-08- 8
From 2008.igem.org
| Projects | Events | Resources | ||
|---|---|---|---|---|
| Sponsors | Experiments | Milestones | Protocols | |
| Notebook (t) | Meetings (t) |
Things we did today
Wetlab work
Restreaked nir+rbs and I14032+rbs constructs
- Grace
Plasmid prep (finished up)
- Grace
- Resuspended plasmid preps in 50 μl TE buffer
- Determined DNA concentrations of plasmids
| Plasmid | DNA Concentration |
|---|---|
| nir | 496.3 ng/μl |
| GFPf | 484.8 ng/μl |
| BB-pRL1383a | 508.0 ng/μl |
Prep for sequencing
- Grace
- 25 μl PCR reactions of nir+rbs, I14032+rbs, slr1, slr2, BB-pRL1383a
- Colony PCRs seem to indicate success
- 25 μl PCR reactions of GFP+tt, GFPf+tt, J33207+tt
- Colony PCR indicates no ligation. Picked new colony, sequence to confirm failure.
- PCR of nir, B0015, B0030, B0034 for resequencing (bad reads last time)
- Gel purified all PCR rxns (we still have a problem with contaminant DNA/shadow bands) and desired bands were extracted from gel
- 2% agarose gel ran at 60v for 2 hours
- Determined DNA concentrations via nanodrop spectrometer
- Prepared samples and sent to CORE Hawaii for sequencing
Construct p+r+g and p+r+s
- Krystle
File:080808resdig.jpg
EtBr stained 2% agarose gel ran at 60V for 60 min. Forty microliters of each digest were loaded.
- Restriction Digest
- nir+B0030, I14032+B0030, J33207 digested with SpeI and PstI
- gfp, gfpfusion, and B0015 digested with XbaI and PstI
- Gel Purified restriction digest
- 2% agarose gel ran at 60 volts for 1.5 hours
- 40ul of the total restriction digest loaded into each well
- gfpfusion cut out from gel
- Restriction Digest
Discussion
Quote of the Day
History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson
