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Team:Lethbridge CCS/Notebook
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<!-- {{#calendar: title=Lethbridge_CCS |year=2008 | month=05}} --> | <!-- {{#calendar: title=Lethbridge_CCS |year=2008 | month=05}} --> | ||
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| + | === 2 Sept 2008 === | ||
| + | |||
| + | (Peter, Paul, Elizabeth, Glenda, Nathan) | ||
| + | |||
| + | * Resuspended our primers (psB1A7 forward and reverse; TetR-GFP forward and reverse) to 100 μM. | ||
| + | * Made 10-fold dilutions of each primer | ||
| + | * Set up and ran PCR on psB1A7 | ||
| + | ** varied concentration of vector (1 x, 1/10 x, 1/100 x) and annealing temperatures (58°C, 62°C, 66°C), giving nine variations; | ||
| + | ** used materials and amounts as per Phusion polymerase kit | ||
| + | |||
| + | - Cycling temperatures and times for PCR | ||
| + | Initial denaturation 98°C 30 sec ___ | ||
| + | Denaturation 98°C 10 sec | | ||
| + | Annealing 58/62/66 °C 30 sec |--- 30 cycles | ||
| + | Extension 72°C 1 min ___| | ||
| + | Final extension 72°C 10 min | ||
| + | Hold 4°C hold | ||
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| + | |||
| + | <!-- [[Image:pic, if needed]] --> | ||
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=== 6 Sept 2008 === | === 6 Sept 2008 === | ||
Revision as of 21:57, 6 September 2008
2 Sept 2008
(Peter, Paul, Elizabeth, Glenda, Nathan)
- Resuspended our primers (psB1A7 forward and reverse; TetR-GFP forward and reverse) to 100 μM.
- Made 10-fold dilutions of each primer
- Set up and ran PCR on psB1A7
- varied concentration of vector (1 x, 1/10 x, 1/100 x) and annealing temperatures (58°C, 62°C, 66°C), giving nine variations;
- used materials and amounts as per Phusion polymerase kit
- Cycling temperatures and times for PCR
Initial denaturation 98°C 30 sec ___
Denaturation 98°C 10 sec |
Annealing 58/62/66 °C 30 sec |--- 30 cycles
Extension 72°C 1 min ___|
Final extension 72°C 10 min
Hold 4°C hold
6 Sept 2008
- Agarose gel purification of pSB1A7 from [date] (Peter, Paul, Marc, Glenda, Nathan)
- conditions
