Complete homology maps of the rabbit (Oryctolagus cuniculus) and human by reciprocal chromosome painting

Fluorescence in situ hybridization (FISH) was used to construct a homology map to analyse the extent of evolutionary conservation of chromosome segments between human and rabbit (Oryctolagus cuniculus, 2n = 44). Chromosome-specific probes were established by bivariate fluorescence activated flow sorting followed by degenerate oligonucleotide-primed PCR (DOP-PCR). Painting of rabbit probes to human chromosomes and vice versa allowed a detailed analysis of the homology between these species. All rabbit chromosome paints, except for the Y paint, hybridized to human chromosomes. All human chromosome paints, except for the Y paint, hybridized to rabbit chromosomes. The results obtained revealed extensive genome conservation between the two species. Rabbit chromosomes 12, 19 and X were found to be completely homologous to human chromosomes 6, 17 and X, respectively. All other human chromosomes were homologous to two or sometimes three rabbit chromosomes. Many conserved chromosome segments found previously in other mammals (e.g. cat, pig, cattle, Indian muntjac) were also found to be conserved in rabbit chromosomes.     

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

Background

The 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.

Results

The 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.

Conclusion

The 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.

     

Response to cecropia cocoons of Mus musculus and two species of Peromyscus

Summary In rural areas cocoons of overwintering Hyalophora cecropia spum close to the ground are opened and the pupae eaten by small mammals, but in urban areas, where cecropia populations are much larger, the cocoons are almost never opened by mammals. Peromyscus leucopus noveboracensis and P. maniculatus bairdii are abundant in rural areas, but are largely replaced by Mus musculus in urban areas. In laboratory experiments P. l. noveboracensis invariably opened cecropia cocoons and ate the pupae. P. m. bairdii were slower to open cecropia cocoons and two of twelve did not open one at all. M. musculus, on the other hand, never opened cecropia cocoons although they gnawed the silk and ate uncovered pupae. Cocoons opened in the laboratory by the two species of Peromyscus cannot be distinguished from each other or from cocoons opened in the field.Part of the Ph. D. dissertation presented by the first author to the Department of Entomology, University of Illinois. This investigation was supported in part by PHS Training Grant no. 2-T01-GM 01076 from General Medical Sciences.     

Comparative genomics: tracking chromosome evolution in the family ursidae using reciprocal chromosome painting

The Ursidae family includes eight species, the karyotype of which diverges somewhat, in both chromosome number and morphology, from that of other families in the order Carnivora. The combination of consensus molecular phylogeny and high-resolution trypsin G-banded karyotype analysis has suggested that ancestral chromosomal fissions and at least two fusion events are associated with the development of the different ursid species. Here, we revisit this hypothesis by hybridizing reciprocal chromosome painting probes derived from the giant panda (Ailuropoda melanoleuca), domestic cat (Felis catus), and man (Homo sapiens) to representative bear species karyotypes. Comparative analysis of the different chromosome segment homologies allowed reconstruction of the genomic composition of a putative ancestral bear karyotype based upon the recognition of 39 chromosome segments defined by painting as the smallest conserved evolutionary unit segments (pSCEUS) among these species. The different pSCEUS combinations occurring among modern bear species support and extend the postulated sequence of chromosomal rearrangements and provide a framework to propose patterns of genome reorganization among carnivores and other mammal radiations.     

Mus and Peromyscus chromosome homology established by FISH with three mouse paint probes

Fluorescence-labeled DNA probes constructed from three whole house mouse (Mus domesticus) chromosomes were hybridized to metaphase spreads from deer mouse (Peromyscus maniculatus) to identify homologies between the species. Mus Chr 7 probe hybridized strongly to the ad-centromeric two-thirds of Peromyscus Chr 1q. Most of Mus 3 probe hybridized principally to two disjunct segments of Peromyscus Chr 3. Mus Chr 9 probe hybridized entirely to the whole Peromyscus Chr 7. Three Peromyscus linkage groups were assigned to chromosomes, based on linkage homology with Mus. The data also are useful in interpretation of chromosomal evolutionary history in myomorphic rodents. Received: 1 December 1998 / Accepted: 17 February 1999     

Reciprocal chromosome painting between three laboratory rodent species

The laboratory mouse (Mus musculus, 2n = 40), the Chinese hamster (Cricetulus griseus, 2n = 22), and the golden (Syrian) hamster (Mesocricetus auratus, 2n = 44) are common laboratory animals, extensively used in biomedical research. In contrast with the mouse genome, which was sequenced and well characterized, the hamster species has been set aside. We constructed a chromosome paint set for the golden hamster, which for the first time allowed us to perform multidirectional chromosome painting between the golden hamster and the mouse and between the two species of hamster. From these data we constructed a detailed comparative chromosome map of the laboratory mouse and the two hamster species. The golden hamster painting probes revealed 25 autosomal segments in the Chinese hamster and 43 in the mouse. Using the Chinese hamster probes, 23 conserved segments were found in the golden hamster karyotype. The mouse probes revealed 42 conserved autosomal segments in the golden hamster karyotype. The two largest chromosomes of the Chinese hamster (1 and 2) are homologous to seven and five chromosomes of the golden hamster, respectively. The golden hamster karyotype can be transformed into the Chinese hamster karyotype by 15 fusions and 3 fissions. Previous reconstructions of the ancestral murid karyotype proposed diploid numbers from 2n = 52 to 2n = 54. By integrating the new multidirectional chromosome painting data presented here with previous comparative genomics data, we can propose that syntenies to mouse Chrs 6 and 16 were both present and to hypothesize a diploid number of 2n = 48 for the ancestral Murinae/Cricetinae karyotype.     

Reproductive failure and maternal-fetal relationship in a Peromyscus species cross.

Mating, fertilization, implantation, prenatal mortality, fetal and placental size, and placental ultrastructure were studied in intraspecific and interspecific crosses involving Peromyscus maniculatus and P. polionotus. Failure to mate was a major factor in interspecific crosses and was much more pronounced in crosses between P. polionotus females and P. maniculatus males than in the reciprocal cross. Failure of implantation following mating, however, was more pronounced in crosses between P. maniculatus females and P. polionotus males. Failure of implanted embryos to survive to term was a factor in crosses between P. polionotus females and P. maniculatus males. Comparison of the placental labyrinth of conceptuses from intraspecific and interspecific crosses revealed no differences at the ultrastructural level. The relationship of these observations to the evolution of isolating mechanisms in mammals and to physiological aspects of the developing maternal-fetal relationship are discussed. A model of placental and fetal size inheritance is presented.     

Comparative chromosome painting in Aotus reveals a highly derived evolution

The genus Aotus represents a highly diverse group with an especially intricate taxonomy. No standard cytogenetic nomenclature for the genus has yet been established. So far, cytogenetic studies have characterized 18 different karyotypes with diploid numbers ranging from 46 to 58 chromosomes. By combining G-banding comparisons and molecular cytogenetic techniques, we were able to describe the most likely pattern of chromosome evolution and phylogenetic position of two Aotus karyomorphs (KMs) from Venezuela: Aotus nancymai (KM3, 2n=54) and Aotus sp. (KM9, 2n=50). All of the proposed Platyrrhini ancestral associations (2/16, 3/21, 5/7, 8/18, 10/16, 14/15) were found in the Aotus KMs studied, except 2/16 and 10/16. In addition, some derived chromosomal associations were also detected in both KMs (1/3, 1/16, 2/12, 2/20, 3/14, 4/15, 5/15, 7/11, 9/15, 9/17, 10/11, and 10/22). Although some of these associations have been found in other New World monkeys, our results suggest that Aotus species have undergone a highly derived chromosomal evolution. The homologies between these two Aotus KMs and human chromosomes were established, indicating that KM3 has a more derived karyotype than KM9 with respect to the ancestral Platyrrhini karyotype.     

RAPD-based genetic linkage map of blueberry derived from a cross between diploid species (Vaccinium darrowi and V. elliottii)

An initial genetic linkage map for blueberry has been constructed from over 70 random amplified polymorphic DNA (RAPD) markers that segregated 11 in a testcross population of 38 plants. The mapping population was derived from a cross between two diploid blueberry plants: the F₁ interspecific hybrid (Vaccinium darrowi Camp x V. elliottii Chapm.) and another V. darrowi plant. The map currently comprises 12 linkage groups (in agreement with the basic blueberry chromosome number) and covers a total genetic distance of over 950cM, with a range of 3–30cM between adjacent markers. The use of such a map for identifying molecular markers linked to genes controlling chilling requirement and cold hardiness is discussed.     

A cosmid clone map derived from a small region of human chromosome 11

We have used cosmid "fingerprinting" to construct an overlapping DNA clone "map" of the human DNA in a mouse/human hybrid cell line, E65-9, that contains about 4 x 10(6) bp, including the H-Ras gene, as its human component. We have additionally used 32P-labeled RNA probes to establish linkage of particular sets of clones, and the final map comprises about 300,000 bp and is contained in three nonoverlapping segments. The reasons for failure to close the gaps by direct probing are discussed. We have developed techniques to search for rare cutting restriction enzyme cleavage sites in large numbers of cloned DNAs and have positioned sites for EagI and BssHII on our clone map. The methods we used are capable of considerable scale-up and are currently being applied to the short arm of human chromosome 11.     

Analysis of reciprocal translocations by chromosome painting: applications and limitations of the technique.

Fluorescent in situ hybridization with chromosome-specific DNA libraries (chromosome painting) is an important new method for assessing chromosome rearrangements. In the research presented in this paper, two familial reciprocal translocations have been studied in the balanced and unbalanced forms, using both traditional G-banding techniques and chromosome painting. Although for each case two chromosomes were involved in the rearrangement, we found that only one chromosome library was suitable for detecting the translocation. These findings illustrate both the potential and the limitations of chromosome painting as a diagnostic tool in cytogenetics.     

Karyotype evolution of eulipotyphla (insectivora): the genome homology of seven sorex species revealed by comparative chromosome painting and banding data

The genus Sorex is one of the most successful genera of Eulipotyphla. Species of this genus are characterized by a striking chromosome variability including XY1Y2 sex chromosome systems and exceptional chromosomal polymorphisms within and between populations. To study chromosomal evolution of the genus in detail, we performed cross-species chromosome painting of 7 Sorex species with S. granarius and S. araneus whole-chromosome probes and found that the tundra shrew S. tundrensis has the most rearranged karyotype among these. We reconstructed robust phylogeny of the genus Sorex based on revealed conserved chromosomal segments and syntenic associations. About 16 rearrangements led to formation of 2 major Palearctic groups after their divergence from the common ancestor: the S. araneus group (10 fusions and 1 fission) and the S. minutus group (5 fusions). Further chromosomal evolution of the 12 species inside the groups, including 5 previously investigated species, was accompanied by multiple reshuffling events: 39 fusions, 20 centromere shifts and 10 fissions. The rate of chromosomal exchanges upon formation of the genus was close to the average rate for eutherians, but increased during recent (about 6-3 million years ago) speciation within Sorex. We propose that a plausible ancestral Sorex karyotype consists of 56 elements. It underwent 20 chromosome rearrangements from the boreoeutherian ancestor, with 14 chromosomes retaining the conserved state. The set of genus-specific chromosome signatures was drawn from the human (HSA)-shrew comparative map (HSA3/12/22, 8/19/3/21, 2/13, 3/18, 11/17, 12/15 and 1/12/22). The syntenic association HSA4/20, that was previously proposed as a common trait of all Eulipotyphla species, is shown here to be an apomorphic trait of S. araneus.     

A novel source of highly specific chromosome painting probes for human karyotype analysis derived from primate homologues

We have established a series of highly specific painting probes for human acrocentric chromosomes. These chromosomes are involved in the formation of the nucleolar organizer region (NOR) and show DNA sequence homologies within their pericentric heterochromatin. To date, these chromosomes have shown considerable cross hybridization in chromosome painting experiments. Our probe set has been established from primate homologues that are not involved in the NOR in that particular species or from species in which highly repetitive sequences have undergone rapid sequence divergence. The new painting probes should be of particular value for automated microscopy, for which highly specific signals are required as they are recorded at low magnification, e.g. when scoring chromosome 21 domains in interphase nuclei. Received: 22 May 1997 / Accepted: 16 June 1997     

Multidirectional chromosome painting reveals a remarkable syntenic homology between the greater galagos and the slow loris

We report on the first reciprocal chromosome painting of lorisoids and humans. The chromosome painting showed a remarkable syntenic homology between Otolemur and Nycticebus. Eight derived syntenic associations of human segments are common to both Otolemur and Nycticebus, indicative of a considerable period of common evolution between the greater galago and the slow loris. Five additional Robertsonian translocations form the slow loris karyotype, while the remaining chromosomes are syntenically equivalent, although some differ in terms of centromere position and heterochromatin additions. Strikingly, the breakpoints of the human chromosomes found fragmented in these two species are apparently identical. Only fissions of homologs to human chromosomes 1 and 15 provide significant evidence of a cytogenetic link between Lemuriformes and Lorisiformes. The association of human chromosomes 7/16 in both lorisoids strongly suggests that this chromosome was present in the ancestral primate genome.     

Sperm chromosome complements from two human reciprocal translocation heterozygotes

Summary Using the hamster oocyte/human sperm fusion technique, we studied sperm chromosome complements in two male reciprocal translocation heterozygotes, 46,XY,t(11;17)(p11.2;q12.3) and 46,XY,t(1;11) (p36.3;q13.1). For the t(11;17) carrier, 202 sperm chromosome complements were obtained, but 18 karyotypes were not included in the segregation data because of multiple breaks and rearrangements. The alternate and adjacent I types, adjacent II, and 31 segregations accounted for 38.6%, 32.1%, 26.6%, and 2.7% of the sperm analyzed from the t(11;17) carrier. A total of 575 sperm chromosome complements was obtained using sperm from the t(1;11) heterozygote, and 27 karyotypes were excluded from the segregation data because of multiple breaks and rearrangements. For the t(1;11) carrier, the alternate and adjacent I types, adjacent II, and 31 segregations were responsible for 31.4%, 42.9%, 15.9%, and 8.0% of the analyzed sperm chromosome complements. Chromosomal abnormalities unrelated to the translocation, particularly the conservative estimate of aneuploidy frequency, were within the range observed in normal men. Hence, there was no evidence for an interchromosomal effect causing meiotic nondisjunction, despite the large sample sizes studied.     

An extraordinarily stable karyotype of the woody Populus species revealed by chromosome painting

The karyotype represents the basic genetic make‐up of a eukaryotic species. Comparative cytogenetic analysis of related species based on individually identified chromosomes has been conducted in only a few plant groups and not yet in woody plants. We have developed a complete set of 19 chromosome painting probes based on the reference genome of the model woody plant Populus trichocarpa. Using sequential fluorescence in situ hybridization we were able to identify all poplar chromosomes in the same metaphase cells, which led to the development of poplar karyotypes based on individually identified chromosomes. We demonstrate that five Populus species, belonging to five different sections within Populus, have maintained a remarkably conserved karyotype. No inter‐chromosomal structural rearrangements were observed on any of the 19 chromosomes among the five species. Thus, the chromosomal synteny in Populus has been remarkably maintained after nearly 14 million years of divergence. We propose that the karyotypes of woody species are more stable than those of herbaceous plants since it may take a longer period of time for woody plants to fix chromosome number or structural variants in natural populations.