title: Generation of nonpolar deletion mutants in Listeria monocytogenes using the "SOEing" method.
abstract: The ability to manipulate chromosomally encoded genes is a fundamental biological tool for the analysis of gene function. Here, we provide in greater depth protocols for the creation of nonpolar unlabeled gene deletions in Listeria (L.) monocytogenes that are facilitated by the splicing overlap extension PCR technique. For mutagenesis in L. monocytogenes, we describe two different plasmid-based approaches, which facilitate the introduction of this spliced amplicon in place of the corresponding segment of chromosomal DNA: the pKSV7 system and the pORI280/pVE6007 system.
authors.affiliation: University of Veterinary Medicine Vienna, Veterinaerplatz 1, 1210, Vienna, Austria, kathrin.rychli@vetmeduni.ac.at.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Rychli
authors.forename: Caitriona M
authors.initials: CM
authors.lastname: Guinane
authors.forename: Karen
authors.initials: K
authors.lastname: Daly
authors.forename: Colin
authors.initials: C
authors.lastname: Hill
authors.forename: Paul D
authors.initials: PD
authors.lastname: Cotter
mesh_terms: D002876:Chromosomes, Bacterial / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods; D018274:Electroporation / Q000379:methods; D017353:Gene Deletion; D005822:Genetic Vectors / Q000235:genetics; D008089:Listeria monocytogenes / Q000235:genetics*; D008088:Listeriosis / Q000382:microbiology; D016296:Mutagenesis; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D014170:Transformation, Genetic