Recombineering: Difference between revisions
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==Protocols== | ==Protocols== | ||
*[[/Lambda red-mediated gene replacement|λ red-mediated gene replacement]] — Datsenko and Wanner method, using plasmids | *[[/Lambda red-mediated gene replacement|λ red-mediated gene replacement]] — Datsenko and Wanner method, using plasmids | ||
*[[ | *[[Sauer:Plasmid-born λRed recombineering using dsDNA|Court lab λRed temperature sensitive system]] — Court method using temperature sensitive plasmids (pSIM5) | ||
==Resources== | ==Resources== |
Revision as of 12:11, 3 December 2007
Protocols
- λ red-mediated gene replacement — Datsenko and Wanner method, using plasmids
- Court lab λRed temperature sensitive system — Court method using temperature sensitive plasmids (pSIM5)
Resources
Recombineering is a method where E-coli undergoes recombination between linear DNA, introduced through electroporation, with circularized DNA already present within the cell.
http://recombineering.ncifcrf.gov/
[math]\displaystyle{ \lambda }[/math] phage exo, bet, and gam genes are integrated into E-coli to demonstrate phage promoted homologous recombination.
Murphy 1998 [math]\displaystyle{ \lambda }[/math] Red-Promoted Gene Replacement
To acheive control over phage recombination gene activity, temperature sensitive [math]\displaystyle{ \lambda }[/math] repressor (cI857) was introduced.
Yu 2000 An efficient recombination system for chromosome engineering in Escherichia coli
Court Lab.
This process is then adapted to E-coli strain DH10B, a Bacteria Artificial Chromosome (BAC)strain. This results in the original recombineering E-coli strain designated DY380 as well as other versions modified to include different selection mechanisms. Lee 2001. A highly efficient Escherichia coli-based chromosome engineering system adapted for recombinogenic targeting and subcloning of BAC DNA
Church Lab.
Methods for generating precise deletions and insertions in the genome of wild-type Escherichia coli: application to open reading frame characterization-
All about the [math]\displaystyle{ \lambda }[/math] phage recombination proteins, exonuclease, [math]\displaystyle{ \beta }[/math], and [math]\displaystyle{ \gamma }[/math]!!
An Exonuclease Induced by phage [math]\displaystyle{ \lambda }[/math]. Little 1967
Roles of Exonuclease and [math]\displaystyle{ \beta }[/math] protein of phage [math]\displaystyle{ \lambda }[/math] in recombination. Carter 1971
[math]\displaystyle{ \beta }[/math] protein of phage [math]\displaystyle{ \lambda }[/math] promotes renaturation of DNA. Kmiec 1981
Pairing activities of [math]\displaystyle{ \beta }[/math] protein in phage [math]\displaystyle{ \lambda }[/math]. Muniyappa 1986
How [math]\displaystyle{ \gamma }[/math] protein specified by phage [math]\displaystyle{ \lambda }[/math] affects recBC enzyme in E. Coli. Karu 1975
[math]\displaystyle{ \gamma }[/math] protein of phage [math]\displaystyle{ \lambda }[/math] inhibits recombination activites of RecBCD enzyme. Murphy 1991