The Genetic Manipulation of Staphylococci Methods and Protocols 1st Edition Jeffrey 9781493931576

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Product details:

• ISBN 13: 9781493931576
• Author: Jeffrey L. Bose

This detailed book contains chapters using unbiased approaches such as chemical and transposon mutagenesis, as well as a protocol for allelic exchange, to make targeted mutants in Staphylococcus, as generating mutants of this bacteria can be accomplished in many ways, depending on the intended changes. Using these methods, the authors have been successful at making mutants that span individual single-nucleotide changes in the chromosome to whole genome mutant libraries. The latter strategy provides a useful tool for high-throughput screening, while single nucleotide changes are an elegant way of teasing apart the importance of single nucleotides in gene expression or to create specific amino acid substitutions to examine protein function. Written for the Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Table of contents:

Understanding Staphylococcal Nomenclature

Restriction-Modification Systems as a Barrier for Genetic Manipulation of Staphylococcus aureus

Complementation Plasmids, Inducible Gene-Expression Systems, and Reporters for Staphylococci

De Novo Assembly of Plasmids Using Yeast Recombinational Cloning

Splicing by Overlap Extension PCR to Obtain Hybrid DNA Products

Method for Preparation and Electroporation of S. aureus and S. epidermidis

Rapid Isolation of DNA from Staphylococcus

Bacteriophage Transduction in Staphylococcus aureus: Broth-Based Method

Bacteriophage Transduction in Staphylococcus aureus

Batch Transduction of Transposon Mutant Libraries for Rapid Phenotype Screening in Staphylococcus

Conjugative Transfer in Staphylococcus aureus

Allelic Exchange

Creation of Staphylococcal Mutant Libraries Using Transposon Tn917

Generation of a Transposon Mutant Library in Staphylococcus aureus and Staphylococcus epidermidis Using Bursa aurealis

Chemical and UV Mutagenesis

Pulsed Field Gel Electrophoresis

RNA-Sequencing of Staphylococcus aureus Messenger RNA

Quantitative Real-Time PCR (qPCR) Workflow for Analyzing Staphylococcus aureus Gene Expression

Electrophoretic Mobility Shift Assays

Rapid Amplification of cDNA Ends for RNA transcript Sequencing in Staphylococcus.

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