Difference between revisions of "Help:Standardization"
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#First off, check your DNA sequence for restriction sites. This is easy using available web programs such as | #First off, check your DNA sequence for restriction sites. This is easy using available web programs such as | ||
| − | #*[http://tools.neb.com/NEBcutter2/index.php NEB cutter] | + | #*[http://tools.neb.com/NEBcutter2/index.php NEB cutter] - by New England Biolabs |
#*[http://wishart.biology.ualberta.ca/PlasMapper/ PlasMapper] - developed by the University of Alberta | #*[http://wishart.biology.ualberta.ca/PlasMapper/ PlasMapper] - developed by the University of Alberta | ||
| − | #Next, locate and remove all of the BioBrick standard restriction sites (XbaI, SpeI, EcoRI, PstI) that are on your sequence. To delete | + | #Next, locate and remove all of the BioBrick standard restriction sites (XbaI, SpeI, EcoRI, PstI) that are on your sequence. To delete sites, there are a few processes you can use: |
| − | #* | + | #*site directed mutagenesis - best for changing or deleting small areas of the genome like the Biobricks standard restriction sites. Openwetware has a nice [http://openwetware.org/wiki/Site-directed_mutagenesis protocol on how to do this]. |
| + | #*lambda red - best for if you have very large (thousands of kb, entire genes) areas for alteration or deletion. This process is laid out in [http://www.pnas.org/cgi/content/full/97/12/6640 Datsenko and Wanner, 2000] | ||
Revision as of 15:12, 16 June 2006
What you've got: Any non-Biobrick-standardized sequence of DNA that does something interesting
What you want:A Biobrick standardized sequence of interesting DNA
So here's how you do it...
- First off, check your DNA sequence for restriction sites. This is easy using available web programs such as
- [http://tools.neb.com/NEBcutter2/index.php NEB cutter] - by New England Biolabs
- [http://wishart.biology.ualberta.ca/PlasMapper/ PlasMapper] - developed by the University of Alberta
- Next, locate and remove all of the BioBrick standard restriction sites (XbaI, SpeI, EcoRI, PstI) that are on your sequence. To delete sites, there are a few processes you can use:
- site directed mutagenesis - best for changing or deleting small areas of the genome like the Biobricks standard restriction sites. Openwetware has a nice [http://openwetware.org/wiki/Site-directed_mutagenesis protocol on how to do this].
- lambda red - best for if you have very large (thousands of kb, entire genes) areas for alteration or deletion. This process is laid out in [http://www.pnas.org/cgi/content/full/97/12/6640 Datsenko and Wanner, 2000]