In situ hybridization

In situ hybridization is a cytochemical technique for localizing specific DNA or RNA sequences within an organism. Applications of in situ hybridization technology are diverse. The most common usage in plant cell biology is localization of a messenger RNA (mRNA) at the light microscopic level. This chapter focuses on in situ hybridization and its applications, such as localization of DNA elements on isolated chromosomes, localization of genes or transcripts at the subcellular level by electron microscopy, and in situ polymerase chain reaction (PCR). There are two major categories of labels and detection systems for in situ hybridization: isotopic and nonisotopic. Both have relative merits, depending on the system used. Isotopic labeling is familiar to most molecular biologists, and standard probe production techniques can be used. However, microautoradiography is an exacting procedure and not always easily mastered. Nonisotopic systems are safer, have potentially higher resolution and sensitivity, and produce probes with longevity. Three types of probes can be used for in situ hybridization: synthetic oligodeoxyribonucleotides, double-stranded DNA probes, and single-stranded RNA probes. Each of these probe types has advantages and disadvantages. © 1995, Academic Press Inc.