Team:Newcastle/Restriction digests
From 2010.igem.org
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==Procedures== | ==Procedures== | ||
- | + | Add the following as mentioned below to make up to a final volume of 20 µl of reaction mix: | |
# 15 µl of DNA/plasmid | # 15 µl of DNA/plasmid | ||
# 1 µl of restriction enzyme 1 | # 1 µl of restriction enzyme 1 | ||
Line 18: | Line 18: | ||
# 2 µl of 10X buffer | # 2 µl of 10X buffer | ||
# 1 µl of water | # 1 µl of water | ||
- | + | Incubate the digestion mixture at 37°C for 3 hours | |
==Notes== | ==Notes== | ||
*No more than 10% of enzyme should be used for a single reaction - glycerol inhibits reaction | *No more than 10% of enzyme should be used for a single reaction - glycerol inhibits reaction | ||
- | *10x buffer must be diluted to 1x i.e. 10% final volume | + | *10x buffer must be diluted to 1x i.e. 10% final volume. |
+ | |||
+ | |||
+ | '''Go back to our [[Team:Newcastle/Protocol list|Protocol List]]''' | ||
{{Team:Newcastle/footer}} | {{Team:Newcastle/footer}} |
Latest revision as of 15:54, 5 August 2010
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Contents |
Restriction digestion
Materials required
- Eppendorf tubes
- Pipettes
- Appropriate DNA/plasmid
- Appropriate restriction enzymes
- 10X buffer
- Water
Procedures
Add the following as mentioned below to make up to a final volume of 20 µl of reaction mix:
- 15 µl of DNA/plasmid
- 1 µl of restriction enzyme 1
- 1 µl of restriction enzyme 2
- 2 µl of 10X buffer
- 1 µl of water
Incubate the digestion mixture at 37°C for 3 hours
Notes
- No more than 10% of enzyme should be used for a single reaction - glycerol inhibits reaction
- 10x buffer must be diluted to 1x i.e. 10% final volume.
Go back to our Protocol List