IGEM:MIT/2007/Protocols: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
m (→PCR) |
(→PCR) |
||
| Line 5: | Line 5: | ||
# Set Reaction Tubes/Plates on Ice | # Set Reaction Tubes/Plates on Ice | ||
# Add the following components in a reaction vessel | # Add the following components in a reaction vessel | ||
* 45 µl Platinum PCR SuperMix | ** 45 µl Platinum PCR SuperMix | ||
* Primers (200 nM final concentration per primer is recommended) | ** Primers (200 nM final concentration per primer is recommended) | ||
* Template DNA solution | ** Template DNA solution | ||
Total volume should be between .5-20 µl. | Total volume should be between .5-20 µl. | ||
# Cap reaction vessel and load into a thermal cycler at | # Cap reaction vessel and load into a thermal cycler at 94°C | ||
# Incubate tubes in a thermal cycler at 94°C for 30 s to 2 min | |||
# Perform 25-35 cycles of PCR amplification | |||
## Denature 94° for 15-30 s | |||
## Anneal 55°C for 15-30 s | |||
## Extend 72°C for 1 min per kb | |||
=== Purification === | === Purification === | ||
Revision as of 10:34, 12 June 2007
Protocols
PCR
Platinum PCR Supermix
- Set Reaction Tubes/Plates on Ice
- Add the following components in a reaction vessel
- 45 µl Platinum PCR SuperMix
- Primers (200 nM final concentration per primer is recommended)
- Template DNA solution
Total volume should be between .5-20 µl.
- Cap reaction vessel and load into a thermal cycler at 94°C
- Incubate tubes in a thermal cycler at 94°C for 30 s to 2 min
- Perform 25-35 cycles of PCR amplification
- Denature 94° for 15-30 s
- Anneal 55°C for 15-30 s
- Extend 72°C for 1 min per kb
