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August/21 August 2009
From 2009.igem.org
transformation
to get new plasmid
- ⑤(RBS+rhlR+terminator+promoter(reg))
- ⑦（RBS＋cinR)
- ⑧（RBS＋luxI)
check the result of transformation
Checked the cell cultures transformed and plated out yesterday (8/10) for colony formation and made an approximate count of the number of colonies.
(plate number)-(location on plate) (no. of colonies)
2-8G 4 2-21F 0 2-6O 2 2-13N <15 ⑤ 0
restriction and digestion
K204009
| Vector | Insert | ||
|---|---|---|---|
| terminater(1-23L) | 10 | ①-3(RBS+LasR) | 12 |
| EcoRI | 1 | EcoRI | 1 |
| XbaI | 1 | SpeI | 1 |
| No.2 | 2 | No.2 | 2 |
| dH2O | 4 | dH2O | 4 |
| total | 20uL | total | 20uL |
K204010
| Vector | Insert | ||
|---|---|---|---|
| RBS(1-2M) | 10 | EpsE+terminater(②-A) | 12 |
| SpeI | 1 | XbaI | 1 |
| PstI | 1 | PstI | 1 |
| No.2 | 2 | No.2 | 2 |
| dH2O | 6 | dH2O | 4 |
| total | 20uL | total | 20uL |
K204011
| Vector | Insert | ||
|---|---|---|---|
| p(tetR)(1-6I) | 10 | RBS+GYP+terminater(1-12M) | 12 |
| SpeI | 1 | XbaI | 1 |
| PstI | 1 | PstI | 1 |
| No.2 | 2 | No.2 | 2 |
| dH2O | 6 | dH2O | 4 |
| total | 20uL | total | 20uL |
↓
37°C, 2hr
↓
gel electrophoresis
gel cut
'8/22
purification by [QIAquick Nucleotide Removal Kit]
↓
ligation
ligation DNA 44 10* buffer 5 ligation 1 total 50uL ↓ 16°C , 4hr
