Molecular cytogenetics has revolutionised the way we presently perceive chromosomes in the animal kingdom. Introduction of a number of new molecular approaches during the 80s and the 90s has enabled us to study the chromosomes at the DNA level. One of the techniques central to this transformation is in situ hybridization (ISH), especially the non-radio- active or the fluorescent version (FISH). In combi- nation with resources generated during the past decade, the technique has been instrumental not only in overcoming the limitations encountered in studying chromosomes through traditional cyto- genetics, but has been a major contributor to a strong and emerging branch – comparative genomics, where ‘chromosome painting’ is a very familiar term. Background Chromosome painting implies FISH mediated high- lighting of chromosomes in metaphase or interphase preparations using composite probes specific for a whole chromosome (whole chromosome paints: WCPs) or a region thereof (partial chromosome paints: PCPs). The probe does not represents a single genomic site but is a cocktail of numerous sites from the originating chromosome, such that the whole chromosome or part thereof is almost completely represented. Hence, the observed signal is in fact an aggregation of several hybridization sites uniformly covering the chromosome, and giving a visual impression of ‘painting’. It is from this term that ‘chromosome painting’ has been coined. There are two terms concurrently floating in the literature, viz., ‘chromosome painting’ and ‘compar- ative chromosome painting’. Though with regards to methodology the two approaches do not differ sig- nificantly from each other, their applications and implications differ widely. In the simplest possible way, the former represents use of the painting probe(s) on chromosomes of the species from which they originate (species specific chromosome painting), while the latter refers to their use across evolutionarily distantly related species. It is relatively difficult to define ‘distantly related’. Hence, for the sake of clarity and convenience in discussion, any use of painting probes on species other than their origin will be referred to as comparative chromosome painting. Application and significance of the two categories of chromosome painting will be discussed at length under individual sections. However, because the probes or ‘paints’ (WCPs or PCPs) are the same for both types of painting, they will be discussed commonly in the following section. Methods in Cell Science 23: 37–55 (2001).  2001 Kluwer Academic Publishers. Printed in the Netherlands. Chromosome painting in farm, pet and wild animal species B. P. Chowdhary 1 & T. Raudsepp 2 1 Department of Veterinary Anatomy and Public Health, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4458, USA; 2 Division of Animal Genetics, The Royal Veterinary and Agriculture University, Grønnegårdsvej 3, 1870 Frederiksberg C, Denmark Abstract. Among the advanced karyotype analysis approaches embraced by animal cytogenetics during the past decade, chromosome painting has had the greatest impact. Generation of chromosome specific paints is considered pivotal to his development. Additionally, ability to use these paints across species (referred to as Zoo-FISH or comparative painting) is undisputedly the most important breakthrough that has contributed to our ability to compare karyotypes of a wide range of evolutionarily highly diverged species. This review introduces the readers to the basic principles underlying chromosome painting, and makes them aware of the tools/resources avail- Key words: Chromosome painting, Comparative genomics, Karyotype evolution, Zoo-FISH able to carry out this research in a variety of animal species. An overview of the current status of com- parative chromosome painting results across closely as well as distantly related species is presented. Findings from different studies show how some seg- mental syntenies are more conserved as compared to others. The comparisons provide insight into the likely constitution of a vertebrate/mammalian ancestral karyotype and help understand some of the intricacies about karyotype evolution. Importance of comparative painting in setting the stage for rapid development of gene maps in a number of econom- ically important species is elaborated.