E-Book Overview
Thirty-one detailed, practical chapters many by the originators themselves present easy-to-follow protocols of in situ hybridization techniques for mapping DNA sequences onto chromosomes, quantitation/localization of RNA in tissues, and detection of virus nucleic acids. The methods serve molecular biologists in most areas of basic and applied research, and in every sort of organism ranging from viruses to Drosophila to humans. The expert contributors provide step-by-step instructions for the preparation of probes for DNA, teleomeric repeats, cDNA sequences, small synthetic oligomers, whole cosmids, mega-size YACs, and chromosome-specific paints from somatic cell hybrids and flow-sorted chromosomes. They also present radioactive and nonradioactive probe labeling procedures with the use of nick translation, random priming, multicolor labeling, PRINS labeling of DNA and RNA, and PCR in situ hybridization. Color illustrations enhance the text. Troubleshooting tips, alternative ways of doing things, and informative explanations of why certain steps are done - aids not usually found in standard journal recipes - guarantee a significant difference in the outcome of experiments.
E-Book Content
CHAPTER 1
Preparation of Human Chromosomal Painting Probes From Somatic Cell Hybrids Nicoletta Antonino
Archidiacono, Forabosco,
Rachele Antonacci, and Mariano Rocchi
1. Introduction Chromosomal in situ suppression (CISS) hybridization with flow-sorted chromosome libraries (FSCL) has provided a very powerful tool in human cytogenetics. This technique allows the painting of specific chromosomes in metaphase spreads and in interphase nuclei (1-4). The usefulness of FSCL is particularly evident, for instance, in identifying the chromosomal origin of de ltovo unbalanced translocations and marker chromosomes, or, more generally, m characterizing those cytogeneti