Koch Lab:Protocols/Unzipping constructs/17mer: Difference between revisions
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The protocol is naturally sub-divided into three parts, followed by a ligation reaction(s). | The protocol is naturally sub-divided into three parts, followed by a ligation reaction(s). | ||
===Anchoring segment=== | ===[[/Anchoring segment|Anchoring segment]]=== | ||
See [[/Anchoring segment|Anchoring segment page]] | |||
===Adapter duplex=== | ===[[/Adapter duplex|Adapter duplex]]=== | ||
See [[/Adapter duplex|Adapter duplex page]] | |||
===Downstream unzipping segment=== | ===[[/Downstream unzipping segment|Downstream unzipping segment]]=== | ||
See [[/Downstream unzipping segment|Downstream unzipping segment page]] | |||
==Ligation, final product formation== | ==Ligation, final product formation== |
Revision as of 21:31, 14 June 2008
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This is a specific protocol for creating a DNA construct that will unzip the "17-mer" sequence from plasmid pCP681 from Craig Peterson lab. Steve used this plasmid in his 2002 Biophysical Journal Paper (PMID 12124289). Originally designed to study nucleosome remodeling, this repetitive DNA is also convenient for studying site-specific DNA-binding proteins via unzipping, because multiple binding sites can be probed from a single-tether. The method here is easily adaptable to other DNA unzipping segments, if a few pitfalls are considered. The protocol below is a more detailed version of that provided in Koch et al. 2002.
Individual segments
The protocol is naturally sub-divided into three parts, followed by a ligation reaction(s).
Anchoring segment
Adapter duplex
Downstream unzipping segment
See Downstream unzipping segment page