From 2007.igem.org

Contents 1 Overview 1.1 Excising the sacB gene from plasmid pJQ200 1.2 Verifying the presence of the lacZ promoter in the plasmid pJQ200 1.3 PCR amplification of the global transcription factors SO1415, SO4157, hlyU

Overview

Plasmid pJQ200

Excising the sacB gene from plasmid pJQ200

The native sacB gene was excised from the plasmid pJQ200 with the restriction enzymes EcoRI and HindIII. In the presence of 10% sucrose, the sacB gene acts as a suicide gene. After ligating a gene into this multiple cloning site, the plasmid pJQ200 should no longer be sucrose sensitive and should retain the gentamicin antibiotic resistance gene. Positive transformants will survive in the presence of gentamicin and sucrose.

Verifying the presence of the lacZ promoter in the plasmid pJQ200

PCR amplification of the global transcription factors SO1415, SO4157, hlyU