NYMU Taipei/Lab Notes/2007 8 27
From 2007.igem.org
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==Phase 1 assembly== | ==Phase 1 assembly== | ||
| - | * ''Objective: pOmpC + (RBS+EYFP)'' | + | * '''Objective: pOmpC + (RBS+EYFP)''' |
*Ligation | *Ligation | ||
**Insert:vector=3:1 | **Insert:vector=3:1 | ||
Revision as of 07:12, 30 August 2007
- plasmid concentration after liquid culture extraction
| part | concentration (ug/ul) |
|---|---|
| pOmpC | 6 |
| EYFP | 30 |
| ECFP | 23 |
| CinR | 15 |
| HSL | 49 |
| pCinHSL | 7 |
| pCinHSLRBS | 12 |
| RBS | 14 |
| D-term | 27 |
Phase 1 assembly
- Objective: pOmpC + (RBS+EYFP)
- Ligation
- Insert:vector=3:1
- Vector: 300ng/ul(2ul pOmpC after enzyme digestion)
- Insert: 100ng/ul(2ul EYFP after enzyme digestion)
- Ligase: 1ul
- Final volume: 10ul
- RT 3h
- Enzyme Digestion (pOmpC)
- pOmpC: 14ul
- SpeI: 1ul
- PstI: 1ul
- 10x buffer: 2, 2ul
- 10x BSA: 2ul
- Total: 20ul
- 37°C, 2h
- Heat inactivation: 65°C 20min
- Enzyme Digestion (EYFP)
- EYFP: 3ul
- XbaI: 1ul
- PstI: 1ul
- 10x buffer: 3, 2ul
- 10x BSA: 2ul
- H2O: 11ul
- Total: 20ul
- 37°C, 2h
- Heat inactivation: 65°C 20min
Others
- liquid culture for tar-EnvZ, pLacIQ in 5 ml LB (Kan:20ug/ml)
Who attend?
- 立翔
- 德銘
- 惇茹
- 香君
- 宥嫺
