Tracking genome organization in rodents by Zoo-FISH

The number of rodent species examined by modern comparative genomic approaches, particularly chromosome painting, is limited. The use of human whole-chromosome painting probes to detect regions of homology in the karyotypes of the rodent index species, the mouse and rat, has been hindered by the highly rearranged nature of their genomes. In contrast, recent studies have demonstrated that non-murid rodents display more conserved genomes, underscoring their suitability for comparative genomic and higher-order systematic studies. Here we provide the first comparative chromosome maps between human and representative rodents of three major rodent lineages Castoridae, Pedetidae and Dipodidae. A comprehensive analysis of these data and those published for Sciuridae show (1) that Castoridae, Pedetidae and Dipodidae form a monophyletic group, and (2) that the European beaver Castor fiber (Castoridae) and the birch mouse Sicista betulina (Dipodidae) are sister species to the exclusion of the springhare Pedetes capensis (Pedetidae), thus resolving an enduring trifurcation in rodent higher-level systematics. Our results together with published data on the Sciuridae allow the formulation of a putative rodent ancestral karyotype (2n = 50) that is thought to comprise the following 26 human chromosomal segments and/or segmental associations: HSA1pq, 1q/10p, 2pq, 2q, 3a, 3b/19p, 3c/21, 4b, 5, 6, 7a, 7b/16p, 8p/4a/8p, 8q, 9/11, 10q, 12a/22a, 12b/22b, 13, 14/15, 16q/19q, 17, 18, 20, X and Y. These findings provide insights into the likely composition of the ancestral rodent karyotype and an improved understanding of placental genome evolution. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Microdissected bovine X chromosome segment delineates homoeologous chromosomal regions in sheep, goat and buffalo

The terminal part of the long arm of the bovine X chromosome (bands Xq41--q43) was microdissected. The DNA thus obtained was PCR amplified, labelled and used as painting probe on cattle, sheep, goat and buffalo chromosomes. In cattle, as expected, distinct hybridization signals were observed on bands Xq41--q43. In sheep and goat, the painting signals were observed on the proximal part of the long arm of the X chromosome, adjacent to the centromere (Xp12--q12). In buffalo, however, the terminal part of the X chromosome involving bands q44--q47 was painted. The findings contribute towards developing a better understanding of the comparative organization of the X chromosome in the four bovidae species. Proposed models of evolutionary rearrangements within the X chromosome of the four species are examined in light of the results obtained.     

Construction of chromosome-specific paints for meta- and submetacentric autosomes and the sex chromosomes in the horse and their use to detect homologous chromosomal segments in the donkey

A pilot study comparing horse and donkey karyotypes on a molecular basis was initiated using the chromosomal microdissection approach. All equine meta- and submetacentric chromosomes, viz. ECA1 to ECA13 and the X and Y chromosomes, were microdissected. The DNA was PCR amplified, non-radioactively labelled and used as probes on equine metaphase chromosomes to confirm their origin. Once tested, the paints were used as probes on donkey metaphase chromosomes to detect homologous chromosomal segments between the two species. The results not only detected conservation of whole chromosome and/or arm synteny between the two karyotypes, but also highlighted varying degrees of rearrangements. The findings also enable deduction of homology between parts of donkey and human karyotypes. In light of the molecular evidence, this study examines the accuracy of the available comparative cytogenetic data between horse and donkey. This revised version was published online in July 2006 with corrections to the Cover Date.     

The molecular basis of chromosome orthologies and sex chromosomal differentiation in palaeognathous birds

Palaeognathous birds (Struthioniformes and Tinamiformes) have morphologically conserved karyotypes and less differentiated ZW sex chromosomes. To delineate interspecific chromosome orthologies in palaeognathous birds we conducted comparative chromosome painting with chicken (Gallus gallus, GGA) chromosome 1–9 and Z chromosome paints (GGA1–9 and GGAZ) for emu, double-wattled cassowary, ostrich, greater rhea, lesser rhea and elegant crested tinamou. All six species showed the same painting patterns: each probe was hybridized to a single pair of chromosomes with the exception that the GGA4 was hybridized to the fourth largest chromosome and a single pair of microchromosomes. The GGAZ was also hybridized to the entire region of the W chromosome, indicating that extensive homology remains between the Z and W chromosomes on the molecular level. Comparative FISH mapping of four Z- and/or W-linked markers, the ACO1/IREBP, ZOV3 and CHD1 genes and the EE0.6 sequence, revealed the presence of a small deletion in the proximal region of the long arm of the W chromosome in greater rhea and lesser rhea. These results suggest that the karyotypes and sex chromosomes of palaeognathous birds are highly conserved not only morphologically, but also at the molecular level; moreover, palaeognathous birds appear to retain the ancestral lineage of avian karyotypes.     

Reciprocal chromosome painting illuminates the history of genome evolution of the domestic cat, dog and human

Domestic cats and dogs are important companion animals and model animals in biomedical research. The cat has a highly conserved karyotype, closely resembling the ancestral karyotype of mammals, while the dog has one of the most extensively rearranged mammalian karyotypes investigated so far. We have constructed the first detailed comparative chromosome map of the domestic dog and cat by reciprocal chromosome painting. Dog paints specific for the 38 autosomes and the X chromosomes delineated 68 conserved chromosomal segments in the cat, while reverse painting of cat probes onto red fox and dog chromosomes revealed 65 conserved segments. Most conserved segments on cat chromosomes also show a high degree of conservation in G-banding patterns compared with their canine counterparts. At least 47 chromosomal fissions (breaks), 25 fusions and one inversion are needed to convert the cat karyotype to that of the dog, confirming that extensive chromosome rearrangements differentiate the karyotypes of the cat and dog. Comparative analysis of the distribution patterns of conserved segments defined by dog paints on cat and human chromosomes has refined the human/cat comparative genome map and, most importantly, has revealed 15 cryptic inversions in seven large chromosomal regions of conserved synteny between humans and cats. This revised version was published online in July 2006 with corrections to the Cover Date.     

Karyotype evolution and phylogenetic relationships of hamsters (Cricetidae, Muroidea, Rodentia) inferred from chromosomal painting and banding comparison

The evolutionary success of rodents of the superfamily Muroidea makes this taxon the most interesting for evolution studies, including study at the chromosomal level. Chromosome-specific painting probes from the Chinese hamster and the Syrian (golden) hamster were used to delimit homologous chromosomal segments among 15 hamster species from eight genera: Allocricetulus, Calomyscus, Cricetulus, Cricetus, Mesocricetus, Peromyscus, Phodopus and Tscherskia (Cricetidae, Muroidea, Rodentia). Based on results of chromosome painting and G-banding, comparative maps between 20 rodent species have been established. The integrated maps demonstrate a high level of karyotype conservation among species in the Cricetus group (Cricetus, Cricetulus, Allocricetulus) with Tscherskia as its sister group. Species within the genera Mesocricetus and Phodopus also show a high degree of chromosomal conservation. Our results substantiate many of the conclusions suggested by other data and strengthen the topology of the Muroidea phylogenetic tree through the inclusion of genome-wide chromosome rearrangements. The derivation of the muroids karyotypes from the putative ancestral state involved centric fusions, fissions, addition of heterochromatic arms and a great number of inversions. Our results provide further insights into the karyotype relationships of all species investigated.     

Refined genome-wide comparative map of the domestic horse, donkey and human based on cross-species chromosome painting: insight into the occasional fertility of mules

We have made a complete set of painting probes for the domestic horse by degenerate oligonucleotide-primed PCR amplification of flow-sorted horse chromosomes. The horse probes, together with a full set of those available for human, were hybridized onto metaphase chromosomes of human, horse and mule. Based on the hybridization results, we have generated genome-wide comparative chromosome maps involving the domestic horse, donkey and human. These maps define the overall distribution and boundaries of evolutionarily conserved chromosomal segments in the three genomes. Our results shed further light on the karyotypic relationships among these species and, in particular, the chromosomal rearrangements that underlie hybrid sterility and the occasional fertility of mules.     

Correspondence of human chromosomes 9, 12, 15, 16, 19 and 20 with donkey chromosomes refines homology between horse and donkey karyotypes

Whole chromosome paints for human (HSA) chromosomes 9, 12, 15 and 20 and arm-specific paints for HSA16p, 19p and 19q were applied on donkey metaphase spreads. All probes, except HSA19p, gave distinct hybridization signals on donkey chromosomes/chromosomal segments. The results show direct segmental homology between human and donkey genomes, and enable refinement of correspondence between donkey and horse karyotypes. Of specific interest is the identification of hitherto unknown correspondence between four equine acrocentric chromosomes (ECA22, 23, 25 and 28) and the donkey chromosomes. Overall, the findings mark the beginning of an ordered study of comparative organization of genomes/karyotypes of the equids, that can shed light on karyotype evolution and ancestral chromosomal condition in the Perissodactyls. This revised version was published online in July 2006 with corrections to the Cover Date.     

Comparative Karyotype of Rat and Mouse Using Bidirectional Chromosome Painting

A comparative karyotype of rat (Rattus norvegicus) and mouse (Mus musculus) based on chromosome G-banding morphology, heterologous chromosome painting results and available gene mapping data is proposed. Whole chromosome painting probes from both species were generated by PARM-PCR amplification of flow sorted chromosomes. Bidirectional chromosome painting identifies 36 segments of syntenic homology and allows us to propose a nearly complete comparative karyotype of mouse and rat (except for RNO 13 p and RNO 19 p12-13). Seven segments completely covered the RNO chromosomes 3, 5, 8, 11, 12, 15 and 18. Eight segments completely covered the MMU chromosomes 3, 4, 6, 7, 9, 12, 18 and 19. The RNO chromosomes 5, 8, 18 show complete homology with the MMU chromosomes 4, 9 and 18, respectively. Bidirectional hybridization results clearly assign 16 segments to subchromosomal regions in both species. Interpretation of the results allows subchromosomal assignment of all the remaining segments apart from seven distributed on chromosomes MMU 15, MMU 10 B2-D3 and MMU 17 E3-E5. The proposed comparative karyotype shows overall agreement with available comparative mapping data. The proposed repartition of syntenic homologous segments between the two species provides useful data for gene-mapping studies. In addition, these results will enable the reconstruction of the karyotype of the Cricetidae and Muridae common ancestor and the definition of the precise rearrangements which have occurred in both mouse and rat lineages during evolution. This revised version was published online in August 2006 with corrections to the Cover Date.     

Evolutionarily conserved cytogenetic changes in hematological malignancies of dogs and humans – man and his best friend share more than companionship

The pathophysiological similarities shared by many forms of human and canine disease, combined with the sophisticated genomic resources now available for the dog, have placed ‘man’s best friend’ in a position of high visibility as a model system for a variety of biomedical concerns, including cancer. The importance of nonrandom cytogenetic abnormalities in human leukemia and lymphoma was recognized over 40 years ago, but the mechanisms of genome reorganization remain incompletely understood. The development of molecular cytogenetics, using fluorescence in situ hybridization (FISH) technology, has played a significant role in our understanding of cancer biology by providing a means for ‘interrogating’ tumor cells for a variety of gross genetic changes in the form of either numerical or structural chromosome aberrations. Here, we have identified cytogenetic abnormalities in naturally occurring canine hematopoietic tumors that are evolutionarily conserved compared with those that are considered characteristic of the corresponding human condition. These data suggest that humans and dogs share an ancestrally retained pathogenetic basis for cancer and that cytogenetic evaluation of canine tumors may provide greater insight into the biology of tumorigenesis.     

Comparative Chromosome Map of the Laboratory Mouse and Chinese Hamster Defined by Reciprocal Chromosome Painting

Cross-species reciprocal chromosome painting was used to determine homologous chromosomal regions between the laboratory mouse and Chinese hamster. When mouse chromosome-specific paints were hybridized to Chinese hamster chromosomes, paints specific for mouse chromosomes 3, 4, 9, 14, 18, 19 and X each painted a single chromosomal region, whilst other mouse paints delineated multiple discrete chromosomal regions. The mouse Y paint produced non-specific signals on Chinese hamster chromosomes. Nineteen mouse autosome paints identified a total of 47 homologous chromosome regions in the genome of the Chinese hamster. Hybridization of Chinese hamster paints to mouse chromosomes not only confirmed the above results, but also identified which of the chromosomal regions of these two species were homologous. In total, 10 Chinese hamster autosomal paints detected 38 homologous autosomal segments in the mouse genome. A comparative chromosome map was established based on these reciprocal chromosome painting patterns. This map forms the basis for exchanging gene mapping information between the species and for studying genome evolution. This revised version was published online in August 2006 with corrections to the Cover Date.     

Isolation of human chromosome 21-specific cosmids and their uses in mapping of cosmid contigs on chromosomal subregions

Summary A cosmid library of 3×10⁵ clones has been constructed from a human x hamster hybrid cell line, 153E9a3, which contains human chromosome 21 (HC21) as the only human chromosome. From 56,500 clones of this library, 229 HC21-specific cosmids have been isolated by their hybridization to total human DNA and by their failure to hybridize to total Chinese hamster DNA. The cosmids isolated were then characterized, of these, 28 cosmids (12.2% of those tested) containedNot1 site(s), and 41 cosmids were localized on the eight subregions of HC21 by differential hybridization withAlu-PCR products obtained from a hybrid mapping panel. The cosmids localized were further integrated into the existing contigs using the end-specific probes of the clone insert. Therefore, they provided useful anchor points for contig mapping and walking.     

Cloning and chromosomal localization of MYO15A to chromosome 5 of the dog (Canis familiaris)

Mutations in the myosin XVA gene (MYO15A) cause congenital non-syndromic deafness in humans and mice. Therefore, the MYO15A gene represents a candidate gene for hereditary hearing loss in dogs. Using a human cDNA to screen a dog BAC library, we isolated a canine BAC clone. Sequencing of the BAC ends confirmed homology to the human gene. To facilitate future linkage studies, we report the physical mapping of the canine MYO15A gene to CFA5q23-q24 by FISH and RH mapping. This revised version was published online in July 2006 with corrections to the Cover Date.     

Highly conserved linkage homology between birds and turtles: Bird and turtle chromosomes are precise counterparts of each other

The karyotypes of birds, turtles and snakes are characterized by two distinct chromosomal components, macrochromosomes and microchromosomes. This close karyological relationship between birds and reptiles has long been a topic of speculation among cytogeneticists and evolutionary biologists; however, there is scarcely any evidence for orthology at the molecular level. To define the conserved chromosome synteny among humans, chickens and reptiles and the process of genome evolution in the amniotes, we constructed comparative cytogenetic maps of the Chinese soft-shelled turtle (Pelodiscus sinensis) and the Japanese four-striped rat snake (Elaphe quadrivirgata) using cDNA clones of reptile functional genes. Homology between the turtle and chicken chromosomes is highly conserved, with the six largest chromosomes being almost equivalent to each other. On the other hand, homology to chicken chromosomes is lower in the snake than in the turtle. Turtle chromosome 6q and snake chromosome 2p represent conserved synteny with the chicken Z chromosome. These results suggest that the avian and turtle genomes have been well conserved during the evolution of the Arcosauria. The avian and snake sex Z chromosomes were derived from different autosomes in a common ancestor, indicating that the causative genes of sex determination may be different between birds and snakes.Matsuda and Nishida-Umehara contributed equally to this work.     

Abnormal chromosomal arrangements in human oocytes

Ninety-one human oocytes, lacking signs of fertilization 50h after insemination in vitro, were investigated cytogeneticallyto assess the frequency and type of chromosomal abnormalities.Chromosome spreading permitted adequate karyotyping in 55 oocytes.Non-determined numerical aberrations occurred with the followingfrequencies: hypohaploidy, 10.9% (6/55), hyperhapJoidy, 14.5%(8/55) and hyperdiploidy, 3.6% (2/55). Total aneuploidy occurredwith a frequency of 29.1% and was observed in oocytes from 30patients. No correlation was found between specific chromosomalaberrations and type of infertility, stimulation treatment orgonadotrophin levels. On the other hand, the frequency of aneuploidywas significantly higher (P < 0.05) in patients >35 yearsof age. Two chromosomal complements (3.6%) had structural rearrangements;one oocyte had both structural and numerical chromosomal abnormalitiesand the other had differently condensed regions on the longarms of three chromosomes from group C. The overall frequencyof chromosomal aberrations was 32.7%. Only two samples containedan additional set of polar body chromosomes. Thirteen oocytespresented sperm chromosomes in an arrested stage of prematurechromosome condensation of the G₁, phase and four oocytes showedasynchronous condensation of pronuclear chromosomes. Finally,it was concluded that the high proportion of chromosomal aberrationsobserved in human oocytes may contribute significantly to abnormalembryonic development in vitro.     

De novo isochromosome 18p in two patients: cytogenetic diagnosis and confirmation by chromosome painting

This report concerns two patients with clinical features typical for tetrasomy 18p syndrome. Chromosomal analysis revealed a male karotype in both cases, with an additional small metacentric marker chromosome, putatively an i(18p). Fluorescent in situ hybridization with a chromosome 18-specific paint confirmed that the marker chromosome consisted of chromosome 18 material in both cases.     

用比较基因组杂交技术(CGH)检测肾癌染色体畸变

目的应用比较基因组杂交技术(CGH)分析原发性肾癌肿瘤组织中染色体异常变化,探讨肾癌细胞遗传物质的改变,揭示肾癌发生发展的内在本质及其与临床特征之间的关系。方法采用CGH技术对12例肾癌组织提取的全基因组DNA进行检测,以了解肾癌全基因组的变化。结果CGH技术检测的12例肾癌标本中均有染色体的畸变(扩增和/或缺失),常见的扩增区是1p、4p、5q、7p、9p、16p,常见的缺失区是3q、4q、6q、9q、14q、18q。结论原发性肾癌存在广泛的遗传物质不平衡现象,肾癌细胞染色体扩增和/或缺失可能是肾癌发生发展的基础。     

Are clinical and biological IVF parameters correlated with chromosomal disorders in early Life: a multicentric study

A multicentric study was carried out to analyse in a large series:(i) the chromosomal status of unfertilized oocytes, (ii) errorsat fertilization and (iii) the chromosomal complement of cleavedembryos. Parameters such as type of sterility, maternal age,stimulation treatment, doses of gonadotrophins administeredand oocyte preincubation time before insemination were studiedin relation to the incidence of chromosome abnormalities. Twenty-sixper cent of the unfertilized oocytes and 29.2% of the embryoshad chromosome anomalies. Maternal age significantly increasedthe rate of aneuploidy in oocytes: 38% in patients over 35 years(versus 24% in younger patients). Fertilization-related abnormalitieswere significant, i.e. 1.6% parthenogenesis and 6.4% polyploidy.Unexplained infertility was correlated with an increase in therate of parthenogenesis (4.2%) when compared with tubal infertility(1.2%). Triploidy was found to be correlated with three parameters.A lower rate of triploidy was observed in the group of couplesreferred because of male sterility (1.9% versus 6.3% for tuba1sterility), in HMG-treated patients (2.4% versus 7% with analoguesof LHRH/HMG) and with a short 2-h preincubation time beforeinsemination (3% versus 7.2% for > 2 h). A general modelfor natural selection against embryos carrying a chromosomeimbalance was proposed.