title: Gene amplification and qRT-PCR.
abstract: This chapter includes methods for the use of the polymerase chain reaction (PCR) with Pseudomonas, and several specific tips for their successful application in this organism. The first part of the chapter includes methods for purifying genomic DNA from, and amplifying genes from, Pseudomonas, in addition to methods which describe how to prepare a cell lysate from Pseudomonas species for colony PCR reactions. The chapter continues with a switch in focus from DNA to RNA, describing methods for RNA isolation from Pseudomonas, cDNA generation, and finally q-RT-PCR to investigate relative changes in gene expression.
authors.affiliation: Department of Life Sciences, MRC Centre for Molecular Bacteriology and Infection (CMBI), Imperial College London, London, UK.
authors.forename: Cerith
authors.initials: C
authors.lastname: Jones
authors.forename: Alain
authors.initials: A
authors.lastname: Filloux
mesh_terms: D003001:Cloning, Molecular; D015169:Colony Count, Microbial; D004269:DNA, Bacterial / Q000302:isolation & purification; D018076:DNA, Complementary / Q000235:genetics; D003851:Deoxyribonucleases / Q000378:metabolism; D005784:Gene Amplification; D005798:Genes, Bacterial / Q000235:genetics; D011549:Pseudomonas / Q000235:genetics* / Q000254:growth & development; D012329:RNA, Bacterial / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
chemical_list: D004269:DNA, Bacterial; D018076:DNA, Complementary; D012329:RNA, Bacterial; D003851:Deoxyribonucleases