NYMU Taipei/Lab Notes/2007 8 27
From 2007.igem.org
Contents |
plasmid concentration after extraction
| part | concentration (ug/ul) | A260 (OD) |
|---|---|---|
| pOmpC | 0.06 | 0.006 |
| EYFP | 0.3 | 0.030 |
| ECFP | 0.23 | 0.023 |
| CinR | 0.15 | 0.015 |
| HSL | 0.49 | 0.049 |
| pCinHSL | 0.07 | 0.007 |
| pCinHSLRBS | 0.12 | 0.012 |
| RBS | 0.14 | 0.014 |
| D-term | 0.27 | 0.027 |
- p.s.1: the sample is diluted to 200X, and one OD is 50 ug/mL
- p.s.2: the calculation of concentration should be OD multiplied only by 10, not by 1,000 (It is a mistake)
- p.s.3: 50 ug/mL = 0.05 ug/uL, and sample is diluted to 200X. Thus the original concentration should be 0.05 ug/uL * 200 = 10 ug/uL for per OD
- p.s.4: [http://www.eppendorfna.com/products/quant_biophotometer_de.asp Eppendorf®Biophotometer]
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
