Team:Wash U/Protocol
From 2009.igem.org
(Difference between revisions)
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:Digestion cuts the DNA at very specific sites with restriction enzymes. This procedure is required for various DNA manipulation techniques including gel electrophoresis and BioBrick assembly. Note: The procedures from this point forward assume that DNA has been amplified and purified. If this is not the case, please read the Polymerase Chain Reaction procedure for amplification and the Mini-Prep procedure for purification. | :Digestion cuts the DNA at very specific sites with restriction enzymes. This procedure is required for various DNA manipulation techniques including gel electrophoresis and BioBrick assembly. Note: The procedures from this point forward assume that DNA has been amplified and purified. If this is not the case, please read the Polymerase Chain Reaction procedure for amplification and the Mini-Prep procedure for purification. | ||
'''Materials''' | '''Materials''' | ||
- | + | * NEBuffer 2 | |
+ | * BSA | ||
+ | * dI H20 | ||
+ | * Upstream, Downstream, and Destination Plasmid parts | ||
+ | * Restriction Enzymes (varies depending on digestion) | ||
'''Procedures''' | '''Procedures''' | ||
# Begin by thawing the upstream, downstream, and destination plasmid parts along with the NEBuffer 2 and BSA. | # Begin by thawing the upstream, downstream, and destination plasmid parts along with the NEBuffer 2 and BSA. | ||
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:Ligation formally joins two or more pieces of DNA together that are already annealed. Note: this procedure requires the products of a successful digestion. | :Ligation formally joins two or more pieces of DNA together that are already annealed. Note: this procedure requires the products of a successful digestion. | ||
'''Materials''' | '''Materials''' | ||
- | + | * DNA Ligase | |
+ | * 10X T4 DNA Ligase Buffer | ||
+ | * dI H20 | ||
+ | * Upstream, Downstream, and Destination Plasmid parts | ||
'''Procedures''' | '''Procedures''' | ||
# Thaw the 10X T4 DNA Ligase Reaction Buffer and mix to dissolve the precipitate. | # Thaw the 10X T4 DNA Ligase Reaction Buffer and mix to dissolve the precipitate. |
Revision as of 15:54, 9 July 2009