From 2008.igem.org
Things we did today
Wetlab work
Verification of transformants
- Grace
EtBr stained 2% agarose gel ran at 95V for 1 hour.
| Construct
| Colonies
|
| nir + rbs+GFP+tt in BBpRL1383a-1
| 2
|
| plac + rbs+GFP+tt in BBpRL1383a-1
| 0
|
| J33207 in pRL1383a (all plates)
| lawn
|
- Colony PCR of transformants
- None of the colonies tested positive for their desired inserts
Transformation
- Margaret
| Construct
| Colonies
|
| lac promoter/rbs(34)+ rep + pSB1A2
| 0
|
| lac promoter/rbs(30)rep/oriV (6) + lac promoter/rbs(34)aada/oriT (A) +pSB1A3
| 113
|
| lac promoter/rbs(30)rep/oriV (6) + lac promoter/rbs(34)aada/oriT (A)
| 141
|
| lac promoter/rbs(30)rep/oriV (6) + lac promoter/rbs(34)aada/oriT (B) +pSB1A3
| 150
|
| oriV (6) + oriT + pSB1A7
| 200
|
| pSB1A7 self (did not de-phosphorylate)
| 50
|
| DH5-a with RP4
| 3
|
| DB3.1 with RP4
| 0
|
| (+) pUC18
| lawn
|
| (-) no plasmid
| 0
|
Construction of GFP secretion devices
- Krystle
- Gel purified restricted nir+ rbs and slr+gfpf
- Dephosphorylated cut pSB1A3 vector
- Ligation
- nir+rbs with slr+gfpf
- lac+rbs with slr+gfpf
- Transformation into home-made competent and commercially competent DH5α cells and plated on LB amp100 plates
Discussion
Quote of the Day
History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson
[http://manoa.hawaii.edu/ 
][http://manoa.hawaii.edu/ovcrge/ 
][http://www.ctahr.hawaii.edu 
]
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