28 Myxococcus xanthus: Expression Analysis

Abstract:

Analysis of gene expression is an important tool for the study of the program involved in development and sporulation of Myxococcus xanthus. A large number of genes have been fused to the β-galactosidase (lacZ) gene, and their expression profiles have been extensively studied using β-galactosidase assays. Recently, two new methods have become available, namely, quantitative PCR (QPCR) and microarray analysis. The QPCR method is feasible only for studying the expression of a limited number of genes (10 to 20 genes), whereas the strength of microarrays is their ability to monitor global gene expression. Microarrays are not suitable or cost-effective for studying gene expression of single genes, but they provide trends of global changes. For QPCR and microarray experiments the RNA must be of good quality; it should therefore be carefully checked for purity and integrity before use. The chapter focuses on the hybridization protocol for amino silane-coated arrays obtained from The Institute for Genomic Research (TIGR). The protocols should be used as starting points for these types of experiments, and some optimization will, especially for microarray experiments, be necessary.