Reciprocal Painting Research Articles

Non-Homologous Sex Chromosomes in Two Geckos (Gekkonidae: Gekkota) with Female Heterogamety

Evaluating homology between the sex chromosomes of different species is an important first step in deducing the origins and evolution of sex-determining mechanisms in a clade. Here, we describe the preparation of Z and W chromosome paints via chromosome microdissection from the Australian marbled gecko (Christinus marmoratus) and their subsequent use in evaluating sex chromosome homology with the ZW chromosomes of the Kwangsi gecko (Gekko hokouensis) from eastern Asia. We show that the ZW sex chromosomes of C. marmoratus and G. hokouensis are not homologous and represent independent origins of female heterogamety within the Gekkonidae. We also show that the C. marmoratus Z and W chromosomes are genetically similar to each other as revealed by C-banding, comparative genomic hybridization, and the reciprocal painting of Z and W chromosome probes. This implies that sex chromosomes in C. marmoratus are at an early stage of differentiation, suggesting a recent origin.

Karyotypic relationships among Equus grevyi, Equus burchelli and domestic horse defined using horse chromosome arm-specific probes

Using laser microdissection we prepared a set of horse chromosome arm-specific probes. Most of the probes were generated from horse chromosomes, some of them were derived from Equus zebra hartmannae. The set of probes were hybridized onto E. grevyi chromosomes in order to establish a genome-wide chromosomal correspondence between this zebra and horse. The use of arm-specific probes provided us with more information on the mutual arrangement of the genomes than we could obtain by means of whole-chromosome paints generated by flow sorting, even if we used reciprocal painting with probe sets from both species. By comparison of our results and results of comparative mapping in E. burchelli, we also established the chromosomal correspondence between E. grevyi and E. burchelli, providing evidence for a very close karyotypic relationship between these two zebra species. Establishment of the comparative map for E. grevyi contributes to the knowledge of the karyotypic phylogeny in the Equidae family.

Phylogenomics of species from four genera of New World monkeys by flow sorting and reciprocal chromosome painting

BackgroundThe taxonomic and phylogenetic relationships of New World monkeys (Platyrrhini) are difficult to distinguish on the basis of morphology and because diagnostic fossils are rare. Recently, molecular data have led to a radical revision of the traditional taxonomy and phylogeny of these primates. Here we examine new hypotheses of platyrrhine evolutionary relationships by reciprocal chromosome painting after chromosome flow sorting of species belonging to four genera of platyrrhines included in the Cebidae family: Callithrix argentata (silvered-marmoset), Cebuella pygmaea (pygmy marmoset), Callimico goeldii (Goeldi's marmoset) and Saimiri sciureus (squirrel monkey). This is the first report of reciprocal painting in marmosets.ResultsThe paints made from chromosome flow sorting of the four platyrrhine monkeys provided from 42 to 45 hybridization signals on human metaphases. The reciprocal painting of monkey probes on human chromosomes revealed that 21 breakpoints are common to all four studied species. There are only three additional breakpoints. A breakpoint on human chromosome 13 was found in Callithrix argentata, Cebuella pygmaea and Callimico goeldii, but not in Saimiri sciureus. There are two additional breakpoints on human chromosome 5: one is specific to squirrel monkeys, and the other to Goeldi's marmoset.ConclusionThe reciprocal painting results support the molecular genomic assemblage of Cebidae. We demonstrated that the five chromosome associations previously hypothesized to phylogenetically link tamarins and marmosets are homologous and represent derived chromosome rearrangements. Four of these derived homologous associations tightly nest Callimico goeldii with marmosets. One derived association 2/15 may place squirrel monkeys within the Cebidae assemblage. An apparently common breakpoint on chromosome 5q33 found in both Saimiri and Aotus nancymae could be evidence of a phylogenetic link between these species. Comparison with previous reports shows that many syntenic associations found in platyrrhines have the same breakpoints and are homologous, derived rearrangements showing that the New World monkeys are a closely related group of species. Our data support the hypothesis that the ancestral karyotype of the Platyrrhini has a diploid number of 2n = 54 and is almost identical to that found today in capuchin monkeys; congruent with a basal position of the Cebidae among platyrrhine families.

Reciprocal painting between humans, De Brazza’s and patas monkeys reveals a major bifurcation in the Cercopithecini phylogenetic tree

We report on reciprocal painting between humans and two Cercopithecini species, Erythrocebus patas (patas monkey) and Cercopithecus neglectus (De Brazza’s monkey). Both human and monkeys chromosome-specific probes were made by degenerate oligonucleotide primed PCR (DOP-PCR) from flow sorted chromosomes. Metaphases of both monkey species were first hybridized with human chromosome-specific probes and then human metaphases were hybridized with chromosome paints from each monkey species. The human paint probes detected 34 homologous segments on the C. neglectus karyotype, while the C. neglectus probes, including the Y, revealed 41 homologous segments on the human karyotype. The probes specific for human chromosomes detected 29 homologous segments in the E. patas karyotype, while the patas monkey probes painted 34 segments on the human karyotype. We tested various hypotheses of Cercopithecini phylogeny and taxonomy developed by morphologists, molecular biologists and cytogeneticists. Our hybridization data confirm that fissions (both Robertsonian and non-Robertsonian) are the main mechanism driving the evolutionary trend in Cercopithecini toward higher diploid numbers and strongly suggest an early phylogenetic bifurcation in Cercopithecini. One branch leads to Cercopithecus neglectus/Cercopithecus wolfi while the other line leads to Erythrocebus patas/Chlorocebus aethiops. Allenopithecus nigroviridis may have diverged prior to this major phylogenetic node.

Reciprocal chromosome painting among human, aardvark, and elephant (superorder Afrotheria) reveals the likely eutherian ancestral karyotype.

The Afrotheria, a supraordinal grouping of mammals whose radiation is rooted in Africa, is strongly supported by DNA sequence data but not by their disparate anatomical features. We have used flow-sorted human, aardvark, and African elephant chromosome painting probes and applied reciprocal painting schemes to representatives of two of the Afrotherian orders, the Tubulidentata (aardvark) and Proboscidea (elephants), in an attempt to shed additional light on the evolutionary affinities of this enigmatic group of mammals. Although we have not yet found any unique cytogenetic signatures that support the monophyly of the Afrotheria, embedded within the aardvark genome we find the strongest evidence yet of a mammalian ancestral karyotype comprising 2n = 44. This karyotype includes nine chromosomes that show complete conserved synteny to those of man, six that show conservation as single chromosome arms or blocks in the human karyotype but that occur on two different chromosomes in the ancestor, and seven neighbor-joining combinations (i.e., the synteny is maintained in the majority of species of the orders studied so far, but which corresponds to two chromosomes in humans). The comparative chromosome maps presented between human and these Afrotherian species provide further insight into mammalian genome organization and comparative genomic data for the Afrotheria, one of the four major evolutionary clades postulated for the Eutheria.

Molecular cytogenetic characterization of the EBV-producing cell line B95-8 (Saguinus oedipus, Platyrrhini) by chromosome sorting and painting.

The cell line B95-8 releases Epstein-Barr virus (EBV) with high titres of transforming activity and is widely used as a model in cancer research and virology. There are, however, controversial reports about the species of origin, cell line stability and karyotype. To address these questions, B95-8 chromosomes were analysed by chromosome sorting and painting by multicolour fluorescence in-situ hybridization. Reciprocal painting was performed between B95-8, 'wildtype' New World monkey and human chromosomes. Saguinus oedipus was revealed as the species of origin. A further five cell-line-specific marker chromosomes, resulting from translocations, deletions and an insertion were found. Although human chromosome 6 or 13 homologues were always involved in these rearrangements, co-hybridization of an EBV-specific DNA probe did not reveal site-specific hybridization to marker chromosomes or at translocation breakpoints. The multicolour probe set described here will be of special value for further evolutionary studies in New World monkeys.

Reciprocal chromosome painting shows that genomic rearrangement between rat and mouse proceeds ten times faster than between humans and cats

Reciprocal chromosome painting between mouse and rat using complete chromosome probe sets of both species permitted us to assign the chromosomal homology between these rodents. The comparative gene mapping data and chromosome painting have a better than 90% correspondence. The reciprocal painting results graphically show that mouse and rat have strikingly different karyotypes. At least 14 translocations have occurred in the 10–20 million years of evolution that separates these two species. The evolutionary rate of chromosome translocations between these two rodents appears to be up to 10 times greater than that found between humans and cats, or between humans and chimpanzees, where over the last 5–6 million years just one translocation has occurred. Outgroup comparison shows that the mouse genome has incorporated at least three times the amount of interchromosomal rearrangements compared to the rat genome. The utility of chromosome painting was also illustrated by the assignment of two new chromosome homologies between rat and mouse unsuspected by gene mapping: between mouse 11 and rat 20 and between mouse 17 and rat 6. We conclude that reciprocal chromosome painting is a powerful method, which can be used with confidence to chart the genome and predict the chromosome location of genes. Reciprocal painting combined with gene mapping data will allow the construction of large-scale comparative chromosome maps between placental mammals and perhaps other animals.