Making delayed-LuxR mutants

From 2009.igem.org

(Difference between revisions)
(New page: ===Experiments=== # luxR gene was mutated by error-prone PCR using Mn2+(Joyce, 1996) and cloned it into a pUC vector. # This library was transformed into XL10-Gold, and the plasmid was mi...)
(Experiments)
 
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# luxR gene was mutated by error-prone PCR using Mn2+(Joyce, 1996) and cloned it into a pUC vector.
# luxR gene was mutated by error-prone PCR using Mn2+(Joyce, 1996) and cloned it into a pUC vector.
# This library was transformed into XL10-Gold, and the plasmid was mini-prepped to get an plasmid library (pUC-[luxR]).
# This library was transformed into XL10-Gold, and the plasmid was mini-prepped to get an plasmid library (pUC-[luxR]).
-
# The plasmid library was transformed into XL1-Blue harboring pAC-plux-gfp, which will glow green when an active LuxR was present.
+
# The plasmid library was transformed into JW1226 harboring pAC-plux-gfp, which will glow green when an active LuxR was present.
# We screened for the fast/dull mutant of LuxR, and collected the clone's plasmid.
# We screened for the fast/dull mutant of LuxR, and collected the clone's plasmid.

Latest revision as of 00:15, 22 October 2009

Experiments

  1. luxR gene was mutated by error-prone PCR using Mn2+(Joyce, 1996) and cloned it into a pUC vector.
  2. This library was transformed into XL10-Gold, and the plasmid was mini-prepped to get an plasmid library (pUC-[luxR]).
  3. The plasmid library was transformed into JW1226 harboring pAC-plux-gfp, which will glow green when an active LuxR was present.
  4. We screened for the fast/dull mutant of LuxR, and collected the clone's plasmid.