1022-7954/02/3808- $27.00 © 2002 MAIK “Nauka /Interperiodica” 0869 Russian Journal of Genetics, Vol. 38, No. 8, 2002, pp. 869–876. Translated from Genetika, Vol. 38, No. 8, 2002, pp. 1034–1042. Original Russian Text Copyright © 2002 by Alkalaeva, Trifonov, Perelman, Graphodatsky. INTRODUCTION Establishing the degree of evolutionary conserva- tion of linkage groups and main tendencies in karyo- type evolution of mammals remains one of the basic problems in comparative cytogenetics. For many years, this task has been resolved by comparing banding pat- terns of differentially painted chromosomes in various species. The data that accumulated in the course of physical genome mapping of humans and certain other species confirmed evolutionary conservation of some linkage groups. It was shown that, the results of com- paring chromosomes may not always be correct when using methods of comparative banding. Moreover, the capacities of physical genome mapping are somewhat limited by the fact that the construction of a saturated genetic map of the genome and involvement of many species from different taxa in this study are time- and resource-consuming. A revolution in comparative cytogenetics was caused by the appearance of an essentially novel tech- nique, comparative chromosome painting or Zoo- FISH, which combines the advantages of cytogenetic and molecular methods. According to this method, chromosome homology regions in various animal spe- cies are revealed via hybridization of DNA library probes of individual chromosomes belonging to one species on metaphase chromosomes of other species. In the course of hybridization, homologous regions can be identified as colored regions (paints). On one hand, comparative chromosome painting reveals homologies between chromosomes at the molecular level, and on the other, allows rapid and efficient comparison of many species. Impediments to this method are the impossibility of estimating the degree of homology between segments (because this method allows only qualitative evaluation of homology) and low resolution limited by painting probe sizes, which prevents the identification of inner inversions and, hence, decreases the number of visible rearrangements. Nevertheless, to date comparative chromosome painting provides a powerful tool for studying the evolution of the mamma- lian karyotype, and soon will be used as such in study- ing the vertebrate evolution in general. SOURCES OF CHROMOSOME-SPECIFIC PROBES Chromosome painting was simultaneously devel- oped by two groups of researchers headed by Pinkel [1] and by Cremer [2]. This technique proved to be suitable for analyzing complicated human chromosomal aberra- tions in clinical diagnostics. Flow-sorted human chro- mosomes isolated by flow cytometry served as original material for painting probes. The first painting probes were chromosome-specific phage libraries. However, a low insertion/vector ratio and low cloning efficiency in phages negatively affected representativeness of the probes. The appearance of DOP–PCR has largely wid- ened the utilization of painting probes [3]. Amplifi- cation of DNAs from flow-sorted chromosomes with a degenerate primer has been used to construct libraries [4]. The libraries constructed by this method are more representative than phage libraries, which allows their use in experiments on heterolo- gous in situ hybridization. However, the large size of fragments isolated upon hybridization, depending on the size of original chro- mosomes, is critical for these probes. Thus, the detec- tion of minor internal rearrangements proved to be problematic. The use of probes containing smaller chromosomal fragments may be helpful in solving this problem. These are either microdissected probes of chromosomal segments or flow-sorted chromosomes of species with the highly rearranged karyotype, such as in gibbon or dog. The application in Zoo-FISH of subchromosomal microdissection libraries and of the libraries obtained from hybrids of somatic cells is limited because due to Comparative Chromosome Painting E. Z. Alkalaeva, V. A. Trifonov, P. L. Perelman, and A. S. Graphodatsky Institute of Cytology and Genetics, Russian Academy of Sciences, Novosibirsk, 630090 Russia; fax: (3832) 33-12-78; e-mail: graf@bionet.nsc.ru Received December 11, 2001; in final form, April 18, 2002 Abstract—The review of the data on comparative chromosomal painting in mammals is presented. The devel- opment of new molecular–cytogenetic methods has resulted in the accumulation of the detailed information on homology of chromosomal segments of more than 50 species from 11 orders. In this review, modern methods of obtaining painting probes are considered in detail, and the basic tendencies of karyotype evolution in differ- ent taxa are discussed. Putative karyotypes of the ancestors of primates, carnivores, and placental mammals are considered.