Microbial gene expression elucidated by selective capture of transcribed sequences (SCOTS)

Abstract

This chapter describes selective capture of transcribed sequences (SCOTS), a technique that combines total ribonucleic acid (RNA) isolation, followed by complementary deoxyribonucleic acid (cDNA) synthesis, hybridization with biotinylated genomic deoxyribonucleic acid (DNA), polymerase chain reaction (PCR) amplification, cloning, and Southern blot hybridization, all of which are commonly used techniques in molecular biology. SCOTS could be used to analyze gene expression of a pathogen present at different sites (i.e., different organs or tissues) within the host. SCOTS could also be used to compare gene expression between mutant and wild-type strains grown under a specific condition. Analysis of microbe-host interactions by SCOTS is a powerful tool for the global analysis of microbial gene expression inside the natural host. Moreover, any potential candidate antigen can be tested for its expression when bacteria are growing in the host cell. Understanding the metabolic pathways used by bacteria growing in host cells could suggest potential targets for development of new antibiotics. The information obtained by SCOTS can lead to identification of specific proteins produced at high levels within antigen-presenting cells, providing candidates for development of effective vaccines against bacterial infections.