Chapter 3 Fluorescent Detection of Nuclear RNA and DNA: Implications for Genome Organization

This chapter discusses the methodology used for the fluorescence detection of individual genes and their nuclear transcripts. The techniques for localizing DNA and RNA are very similar, but with some key differences. It is often necessary to distinguish rigorously between hybridization to DNA versus RNA; hence, it is useful to consider both protocols and their respective differences. The chapter also describes the current technical protocols and illustrates some recent and potential applications in the study of nuclear and chromosome organization. In situ hybridization is a powerful and versatile methodological approach, which makes it possible to visualize specific nucleic acid sequences directly within the morphological context of individual cells, nuclei, and chromosomes. The fluorescence detection technological advances allow for sensitive, high-resolution detection of a single-copy DNA sequence in interphase or metaphase cells, and RNA sequences in the nucleus and cytoplasm. This technique has numerous applications in both biomedical research and clinical diagnostics. It has a large impact on the investigation of genome organization in chromosomes and on the functional state within the interphase nucleus.