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.
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Acknowledgement
Thanks to Dr. Helga Mikkelsen for useful discussion of RNA extraction and q-RT-PCR protocols. Cerith Jones is supported by a PhD fellowship from the Biotechnology and Biological Sciences Research Council (BBSRC). Alain Filloux laboratory is supported by grants from the Wellcome Trust, BBSRC, and the Medical research Council (MRC).
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© 2014 Springer Science+Business Media New York
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Jones, C., Filloux, A. (2014). Gene Amplification and qRT-PCR. In: Filloux, A., Ramos, JL. (eds) Pseudomonas Methods and Protocols. Methods in Molecular Biology, vol 1149. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-0473-0_35
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DOI: https://doi.org/10.1007/978-1-4939-0473-0_35
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-0472-3
Online ISBN: 978-1-4939-0473-0
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