Physical mapping of 20 unmapped fragments of the btau_4.0 genome assembly in cattle, sheep and river buffalo.

The recent advances in sequencing technology and bioinformatics have revolutionized genomic research, making the decoding of the genome an easier task. Genome sequences are currently available for many species, including cattle, sheep and river buffalo. The available reference genomes are very accurate, and they represent the best possible order of loci at this time. In cattle, despite the great accuracy achieved, a part of the genome has been sequenced but not yet assembled: these genome fragments are called unmapped fragments. In the present study, 20 unmapped fragments belonging to the Btau_4.0 reference genome have been mapped by FISH in cattle (Bos taurus, 2n = 60), sheep (Ovis aries, 2n = 54) and river buffalo (Bubalus bubalis, 2n = 50). Our results confirm the accuracy of the available reference genome, though there are some discrepancies between the expected localization and the observed localization. Moreover, the available data in the literature regarding genomic homologies between cattle, sheep and river buffalo are confirmed. Finally, the results presented here suggest that FISH was, and still is, a useful technology to validate the data produced by genome sequencing programs.

Cytogenetic elaboration of a novel reciprocal translocation in sheep.

Reciprocal translocations represent one of the most common structural chromosomal rearrangements observed in both humans and domestic animals. In these translocations, the balanced forms are most frequent but may remain undetected because the carriers show a normal phenotype. For this reason, routine cytogenetic analysis of domestic animals should necessarily rely on banded karyotypes. In fact, during a screening analysis, carried out on phenotypically normal young sheep (Ovis aries, OAR, 2n = 54) from Laticauda-Comisana hybrids, a new structural rearrangement was detected. Two abnormal acrocentric chromosomes (the smallest and the largest one) were found in all metaphases of this carrier animal, suggesting the presence of a reciprocal translocation (rcp). CBA and RBA banding were performed in order to characterize the translocation, and FISH with chromosome-specific BAC probes and telomere probes was applied to confirm the cytogenetic data. The translocation was classified as rcp(4q;12q)(q13;q25).

Molecular characterization of Xp chromosome deletion in a fertile cow.

A young cow of the Marchigiana breed (central Italy) with normal body conformation and external genitalia underwent routine cytogenetic analyses prior to its use for reproduction. After normal chromosome staining, only one X chromosome was observed with a normal diploid number (2n = 60) in all 200 studied cells. Subsequent cytogenetic analyses by using both CBA- and RBA-banding techniques evidenced that almost all the p arms of the other X chromosome was lacking. Detailed FISH-mapping analyses with BAC covering this Xp arm region demonstrated that this large chromosome region was deleted. RBA-banding showed that the deleted X was late replicating. CGH array analysis evidenced that deletion involves the Xp arm from the telomere to around 39.5 Mb, referring to the BosTau6 cattle genome assembly. This abnormality deletes about 40 Mb of the X chromosome sequence, but, despite the large number of genes deleted, none of them are programmed to escape from inactivation. This can explain the normal phenotype of the female which is actually pregnant. Finally, we evidenced, by analysis of an SNP mapped to the deleted region (SNP rs29024121), that the only normal (e.g. nondeleted) X chromosome present derives from the father. Hence, the deletion has a maternal origin.

Advanced comparative cytogenetic analysis of X chromosomes in river buffalo, cattle, sheep, and human.

Based on a recently generated comprehensive gene map for Ovis aries chromosome X (OARX) with an approximately even locus distribution, we assigned selected bacterial artificial chromosome (BAC) probes corresponding to these OARX loci to Bubalus bubalis (BBU) and Bos taurus (BTA) by comparative fluorescence in-situ hybridization (FISH) to improve cytogenetically the X chromosome maps in these species. Twenty-five added loci in BBUX and BTAX, respectively, contribute to a more detailed description of the cytogenetic organization of these chromosomes. Further seven loci were identified in OARX and two DNA probes were assigned to X and Y chromosomes in river buffalo, cattle, and sheep, respectively, and thus identified loci in the pseudoautosomal region. The additional assignments double the number of cytogenetic loci in BBUX and increase their number in BTAX and OARX. The larger quantity of cytogenetic anchors allows a more precise morphological comparison of bovid X chromosomes among each other and with the Homo sapiens (HSA) X chromosome. The anchor loci confirm and refine syntenic fragments in HSAX and identify several evolutionary breakpoints between the compared chromosomes. The cytogenetic assignments in BBUX, BTAX, and OARX represent useable anchors for the ongoing genome sequence assembly in Bovidae.

Chromosome fragility in river buffalo cows exposed to dioxins.

Fifty river buffalo (Bubalus bubalis, 2n = 50) cows reared in two different provinces of Campania (southern Italy) underwent cytogenetic investigations to ascertain possible differences in their chromosome stability. One group (Caserta province) was under legal sequestration due to the presence in the milk mass of higher mean values of dioxins [21.79 pg/g of fat as sum of polychloro-dibenzo-dioxins (PCDDs), polychloro-dibenzo-furans (PCDFs) and dioxin-like polychlorobiphenyls (DL-PCBs)] than both those permitted (6.0 pg/g of fat as WHO-TEQ) and those (1.3 pg/g of fat as WHO-TEQ) observed in the control group raised in Salerno province. Two types of peripheral blood cell cultures were performed: without (normal cultures for the chromosome abnormality (CA) test: chromatid breaks, chromosome breaks, fragments) and with the addition of BrdU for the sister chromatid exchange (SCE) test). The CA test revealed a significantly (P < 0.01) higher chromosome fragility in the exposed cows compared to the control. Indeed, mean values of CA/cell were 1.26 ± 1.15 in exposed cows and 0.37 ± 0.71 in the control. Mean SCE was higher in exposed cows (8.50 ± 3.35) than that (8.29 ± 3.51) found in the control but the difference was not significant. Comparison within the same group of cows at first (FL) and multiple (ML) lactations revealed significantly (P < 0.01) higher mean values of CA/cell in exposed ML-cows vs FL-cows while no statistical differences were found between ML-cows and FL-cows in the control farm. By contrast, significantly (P < 0.01) higher mean values of SCE were found in both groups of FL-cows versus ML-cows. Comparisons with other previous studied species (sheep and cattle) were also performed.

XX SRY-negative true hermaphrodism in two dogs: clinical, morphological, genetic and cytogenetic studies.

This work aimed at giving a deeper insight into peculiar cases of intersexuality occurring in dogs and known as XX true hermaphrodism due to the existence of both testicular and ovarian tissue in one or both gonads in the presence of an XX chromosome constitution. Clinical, histological and genetic approaches were used in the study of an 8-month-old Cocker Spaniel dog and a 3-year-old mixed-breed Pitbull, both showing a female phenotype, clitoromegaly and male behavior. A normal female karyotype (2n = 78,XX) was noticed, and polymerase chain reaction failed to detect SRY in genomic DNA obtained from peripheral blood lymphocytes of both dogs. The reproductive tract was removed by standard ovariohysterectomy and processed for histology. Thereafter, a normal female phenotype was reconstructed by vaginoplasty. Histological examination revealed bilateral ovotestis in both cases: the gonads showed immature testicular parenchyma containing seminiferous tubules, Sertoli and Leydig cells, but no signs of spermatogenesis, together with differently developed ovarian follicles containing oocytes. In the ovotestes, steroidogenesis was detected by P450c17-immunoreactivity in Leydig cells as well as in theca cells, whereas no MIS-immunoreactivity was shown by the Sertoli cells. Genital tracts of Wolffian and Müllerian origin co-existed in both subjects. Both dogs belong to the very rare cases in which testicular tissue develops in the absence of the key gene, SRY. Up to date very few genetic events have been associated with this abnormal sexual differentiation: SOX9 over-expression and RSPO1 mutation. Nevertheless, neither of them has been found in these dogs.

A new and unusual reciprocal translocation in cattle: rcp(11;25)(q11;q14-21).

A new and unusual reciprocal translocation was detected in a heifer of the Agerolese cattle breed during a routine cytogenetic screening carried out on 13 animals (2 males and 11 females) kept at the ConSDABI Conservation Center in Benevento (Southern Italy). The 13 animals investigated had a normal karyotype except for a 1-year-old female, which carried one autosome smaller than the smallest normal bovine autosomes. This small autosome showed very little C-banding in comparison to the other autosomes, while another medium-sized autosome showed 2 distinct and prominent C-bands. RBA-banding and karyotype analysis revealed that these 2 chromosomes were the result of a reciprocal translocation between chromosomes 11 and 25. FISH analysis with BAC142G06 mapping to the proximal (subcentromeric) region of both BTA25 and der11, BAC513H08 (ELN) mapping to BTA25q22dist and der25, and BAC533C11 mapping to the proximal region of BTA11 and der11 confirmed the localization of the breakpoints on band q11 (centromere) of chromosome 11 and q14-21 of chromosome 25. Ag-NOR and sequential RBA/Ag-NOR techniques detected the presence of NORs on both BTA11 and BTA25 and both der11 and der25. To our knowledge, this is the first report of a reciprocal translocation event in cattle with the breakpoint located in the centromeric region.

Extended cytogenetic maps of sheep chromosome 1 and their cattle and river buffalo homoeologues: comparison with the OAR1 RH map and human chromosomes 2, 3, 21 and 1q.

Cytogenetic maps are useful tools for several applications, such as the physical anchoring of linkage and RH maps or genome sequence contigs to specific chromosome regions or the analysis of chromosome rearrangements. Recently, a detailed RH map was reported in OAR1. In the present study, we selected 38 markers equally distributed in this RH map for identification of ovine genomic DNA clones within the ovine BAC library CHORI-243 using the virtual sheep genome browser and performed FISH mapping for both comparison of OAR1 and homoeologous chromosomes BBU1q-BBU6 and BTA1-BTA3 and considerably extending the cytogenetic maps of the involved species-specific chromosomes. Comparison of the resulting maps with human-identified homology with HSA2q, HSA3, HSA21 and HSA1q reveals complex chromosome rearrangements differentiating human and bovid chromosomes. In addition, we identified 2 new small human segments from HSA2q and HSA3q conserved in the telomeric regions of OAR1p and homoeologous chromosome regions of BTA3 and BBU6, and OAR1q, respectively. Evaluation of the present OAR1 cytogenetic map and the OAR1 RH map supports previous RH assignments with 2 main exceptions. The 2 loci BMS4011 and CL638002 occupy inverted positions in these 2 maps.

Chromosome fragility in dairy cows exposed to dioxins and dioxin-like PCBs.

In this study, we compared cross-bred dairy cows in the Susa Valley (Piedmont, northern Italy), reared either near a high-temperature steel production plant (Farms A and B) or in an industry-free area (control). Exposed cows (n = 36) were selected based on mean bulk milk toxic equivalent values of polychlorodibenzodioxins (PCDDs) and dioxin-like (DL) polychlorobiphenyls (PCBs) and polychlorodibenzofurans (PCDFs) equal to 18.56 pg/g fat and 8.56 pg/g of fat in dairy cows from Farms A and B, respectively, exceeding both those permitted by the legislation in force (6 pg/g fat PCDDs and DL-PCDFs/PCBs), and those measured in dairy cows (n = 19) of the farm used as control (1.75 pg/g of fat PCDDs and DL-PCDFs/PCBs). Two types of peripheral blood cell cultures were performed: without (normal cultures for the chromosome abnormality (CA)-test: gaps, chromatid breaks, chromosome breaks and fragments) and with addition of bromodeoxyuridine [for the sister chromatid exchange (SCE)-test]. Both tests revealed a significant (P ≤ 0.05) higher chromosome fragility in the exposed cattle compared to controls: CA/cell mean values (without gaps) were 0.65 ± 0.91, 0.51 ± 0.81 and 0.13 ± 0.39 in Farms A, B and controls, respectively, while SCE/cell mean values were 7.00 ± 2.88, 6.39 ± 2.80 and 5.29 ± 2.51. Although the role of other pollutants (e.g. heavy metals) in the genesis of the recorded chromosome alterations cannot be ruled out, our results confirm the findings of previous research into dioxin-exposed sheep.

A rare case of centric fission and fusion in a river buffalo (Bubalus bubalis, 2n = 50) cow with reduced fertility.

A 5-year-old river buffalo cow underwent cytogenetic investigation since it had only one male offspring, apparently with normal body constitution, which died one month after birth. The female carrier had normal body conformation and internal sex adducts, as revealed by rectal palpation performed by a specialist veterinary practitioner. The cow was found to carry a complex and rare chromosome abnormality. Indeed, a centric fission of one river buffalo (BBU) chromosome 1 with a subsequent (or simultaneous) centric fusion of BBU1p with BBU23 was revealed by both RBA-banding and specific molecular markers of BBU1p (DEFB1) and BBU23 (ACTA2). CBA-banding revealed a pale, very small C-band in the der1 (BBU1q) and a prominent C-band on the new biarmed chromosome originated by rob(1p;23). Both telomeric probes and AgNOR staining confirmed the Robertsonian translocation (rob), both FITC-signals and the NORs (BBU23) being telomerically located. Furthermore, telomeric signals on der1 (BBU1q) indicate that these 2 chromosomal events may be the result of a reciprocal translocation which occurred between BBU1 and BBU23.

Cytogenetic and genetic studies in a hypospadic horse (Equus caballus, 2n = 64).

A 4-year-old male horse of Friesian breed with normal body conformation, development and libido, and showing an evident ventral penis deviation with hypospadias, underwent both cytogenetic and genetic investigation. Although the karyotype showed normal male arrangement (2n = 64,XY), one telomere of horse (ECA) chromosome 1 was shorter than both the other one and those of a normal horse (control), as revealed by CBA- and RBA-banding, and by Ag-NOR and FISH-mapping techniques using telomere PNA probes. Genetic investigation of the SRY and MAMLD1 coding sequences revealed a normal SRY sequence and a mutation in the MAMLD1 gene sequence: a homozygous change (C>A) was found, leading to the synthesis of an isoleucine, instead of a leucine. Although it is difficult to find a strict correlation between hypospadias and the genetic defects revealed by this investigation, this study is the first to be performed in a hypospadic horse using both cytogenetic and genetic investigation.

Chromosomal assignment of R-spondin genes in the donkey (Equus asinus, 2n = 62).

R-spondins constitute a recently discovered small family of growth factors, and the evidence of their role in several developmental pathways is growing fast. In this work we describe the chromosomal location of the four RSPO genes in the donkey. Using horse BACs, we localized RSPO1 on EAS 5q23, RSPO2 on EAS 12q13, RSPO3 on EAS 24q26, and RSPO4 on EAS 15p13. Moreover, RSPO2, RSPO3, and RSPO4 are the first genes mapped on donkey chromosomes 12, 24, and 15, respectively.

Genomic characterization and chromosomal mapping of 5 river buffalo skeletal muscle differentiation master genes.

River buffalo (Bubalus bubalis, 2n = 50, BBU) is a species of economic relevance in a number of countries. This species shows a very peculiar biology and a great capacity for environmental adaptation. There has been an increasing economic interest as well as a growing demand for a more detailed knowledge of molecular features in this species. From this perspective we report a genomic, transcriptional and cytogenetic analysis of 5 master genes involved in skeletal muscle development. Of these 5 genes, MYOD1, MYF5, MYF6 and MYOG belong to the basic helix-loop helix protein family while MSTN belongs to the TNF-B protein family. In mammals, these genes are involved in the early stages of skeletal muscle differentiation, development and regeneration. These pivotal biological functions are finely regulated in a tissue- and temporal-specific manner. We used a comparative genomic approach to obtain the buffalo specific sequences of MYOD1 and MYF6. The nucleotide sequence similarity and the protein domain conservation of the newly obtained sequences are analysed with respect to bovine and other mammalian species showing sequence similarity. The presence of a polymorphism in MYOD1 coding sequence is described and its possible effect discussed. Using a quantitative PCR approach, we compared the level of the 5 transcripts in adult and fetal muscle. These genes were physically localised on river buffalo R-banded chromosomes by FISH using bovine genomic BAC-clones. Here, we present a genomic and cytogenetic analysis which could offer a background to better characterise the buffalo genes involved in muscle function and which may be responsible for buffalo-specific meat features.

Frequency of aneuploidy in in vitro-matured MII oocytes and corresponding first polar bodies in two dairy cattle (Bos taurus) breeds as determined by dual-color fluorescent in situ hybridization.

The current study was undertaken to investigate the aneuploidy rates in in vitro-matured meiosis II (MII) oocytes and corresponding first polar bodies in two dairy cattle (Bos taurus) breeds by using dual-color fluorescent in situ hybridization (FISH). A total of 159 and 144 in vitro-matured MII oocytes of the Italian Friesian and Italian Brown breeds, respectively, were obtained according to the standard methods and analyzed by FISH using "Xcen" and "5" chromosome-specific painting probes, produced by chromosome microdissection and Degenerate Oligonucleotide Primer- Polymerase Chain Reaction (DOP-PCR). Oocytes with unreduced chromosome number were 10.1% and 16.7% in the two breeds, respectively. To avoid bias due to possible artifacts, the aneuploidy rates were determined by analyzing only oocytes with the corresponding polar bodies. In the Italian Friesian, 100 of 143 (69.9%) secondary MII oocytes showed clear MII plates with corresponding first polar bodies and were scored for aneuploidy detection; one oocyte was "nullisomic" for chromosome X (1.0%) and one "disomic" for chromosome 5 (1.0%). In the Italian Brown, 100 of 120 (83.3%) MII oocytes with corresponding first polar bodies were analyzed; one oocyte was nullisomic (1.0%) and one was disomic (1.0%), both for chromosome 5. Totally, 303 oocytes were analyzed, 40 of which showed an unreduced chromosome complement (13.2%); of 200 MII oocytes with the corresponding first polar bodies, the aneuploidy rate (nullisomy+disomy) for the two chromosomes scored was 2%. Assuming that each chromosome is equally involved in aneuploidy, it results that in cattle oocytes matured in vitro, at least 30% of the oocytes (1x30 haploid chromosomes) should be aneuploid. Premature separation of sister chromatids (PSSC) was also observed in 2% of the oocytes in the Italian Friesian breed involving chromosome 5 and in 1% of the Italian Brown breed involving the X chromosome. Estimation of the "baseline" level of aneuploidy in the in vitro-matured oocytes of the various domestic animal species and breeds is, to our opinion, a useful reference for improving the in vitro production of embryos as well as for monitoring future trends of the reproductive health of the species/breeds engaged in zootechnical productions, especially in relation to management errors and environmental hazards.

Comparative mapping and genomic annotation of the bovine oncosuppressor gene WWOX.

WWOX (WW domain-containing oxidoreductase) is the gene mapping at FRA16D HSA16q23.1, the second most active common fragile site in the human genome. In this study we characterized at a detailed molecular level WWOX in the bovine genome. First, we sequenced cDNA from various tissues and obtained evidence in support of a 9-exon structure for the gene, similar to the human gene. Then, we recovered BACs using exon tags and annotated the gene to a >1-Mb genomic region of BTA18 using the Btau 4.0 genome assembly as a reference, thus resolving an issue related to exon 9, which is not included in the genomic annotation of the gene in the Entrez database. Finally, BACs spanning WWOX were used as FISH probes to obtain comparative mapping of the gene in Bos taurus, Bubalus bubalis, Ovis aries and Capra hircus to BTA18q12.1, BBU18q13, OAR14q12.1 and CHI18q12.1, respectively. Our data show that the chromosomal location of WWOX is conserved between man and 4 major domesticated species. Moreover, the annotation of the bovine gene also suggests a highly conserved genomic arrangement, including number and size of introns.

X-Y sperm aneuploidy in 2 cattle (Bos taurus) breeds as determined by dual color fluorescent in situ hybridization (FISH).

The present study was undertaken to investigate aneuploidy rates in the sperm populations of 2 cattle (Bos taurus) breeds by using dual color fluorescent in situ hybridization (FISH) with Xcen and Y chromosome-specific painting probes, obtained by chromosome microdissection and DOP-PCR. Frozen semen from 10 Italian Friesian and 10 Italian Brown testing bulls was used for the investigation. For each bull, more than 5,000 sperm were analyzed, for a total of 52,586 and 51,342 sperm cells for the 2 breeds, respectively. The present study revealed - in both breeds - a preponderance of the Y-bearing sperm compared to the X-bearing sperm. Within each breed, a statistically significant variation in the various classes of aneuploidy (XX, YY and XY) was found: differences were found in the Friesian breed among the 3 diploidy classes, and in the Brown breed, among the 3 disomy classes (p < 0.05) as well as among the 3 diploidy classes (p < 0.01). However, the 2 breeds did not differ significantly in the overall mean rates of X-Y aneuploidy (disomy + diploidy) which amounts to 0.162% in the Italian Friesian and 0.142% in the Italian Brown. When meiosis I (MI) and II (MII) errors were compared, statistically significant differences (p < 0.01) were found in the disomy classes and in both breeds, whereas the differences between diploidy classes were not significant. Compared to humans, a lower level of aneuploidy has been found in the domestic species analyzed so far. The present study contributes to the establishment of a baseline level of aneuploidy in the sperm populations of 2 cattle breeds which could be used for monitoring future trends of reproductive health, especially in relation to environmental changes and mutagens.

The 450-band resolution G- and R-banded standard karyotype of the donkey (Equus asinus, 2n = 62).

Donkey chromosomes were earlier characterized separately by C-, G- and R-banding techniques. However, direct comparisons between G- and R-banding patterns have still not been carried out in this species. The present study reports this comparison at the 450-band level by using replication G- and R-banding patterns. Two sets of synchronized lymphocyte cultures were set up to obtain early (GBA+CBA-banding) and late (RBA-banding) BrdU incorporation. Slides were stained with acridine orange and observed under a fluorescence microscope. Reverse GBA+CBA- and RBA-banded karyotypes at the 450-band level were constructed. To verify G- and R-banding patterns in some acrocentric chromosomes, sequential GBA+CBA/Ag-NORs and RBA/Ag-NORs were also performed. The results of CBA-banding patterns obtained in 12 animals from 2 breeds showed a pronounced polymorphism of heterochromatin, especially in EAS1q-prox. Ideogrammatic representations of G- and R-banded karyotypes were constructed using only one common G- and R-banding nomenclature. In the present study both G- and R-banding patterns and relative ideograms are presented as standard karyotype for this species at the 450-band level.

Copy number variation of testis-specific protein, Y-encoded (TSPY) in 14 different breeds of cattle (Bos taurus).

Multi-copied gene families are prevalent in mammalian genomes, especially within the Y chromosome. Testis specific protein Y-encoded (TSPY) is present in variable copy number in many mammalian species. Previous studies have estimated that TSPY ranges from 50-200 copies in cattle. To examine TSPY localization on the Y chromosome we employed fluorescence in situ hybridization (FISH) and fiber-FISH. The results show a strong signal on the short arm of the Y chromosome (Yp). To investigate TSPY copy number we used relative real-time polymerase chain reaction (PCR) to analyze the DNA of 14 different cattle breeds. Variation both within and between breeds was observed. All breeds show significant variation in TSPY copy number among individual members. Brown Swiss (161 copies, CI = 133-195) had higher average levels of TSPY and Western Fjord Cattle (63 copies, CI = 45-86) had lower levels than some breeds. Overall, however, most breeds had a similar average TSPY copy number. The pooled average was 94 copies (CI = 88-100). The significance of the TSPY array remains uncertain, but as the function of TSPY is unraveled the purpose of the array may become clearer.

FISH-mapping comparison between river buffalo chromosome 7 and sheep chromosome 6: assignment of new loci and comparison with HSA4.

Synchronized peripheral blood lymphocytes from both river buffalo (BBU) and sheep (OAR) were treated for late incorporation of both BrdU and H-33258 to obtain R-banded preparations to be used for FISH-mapping. Ovine BAC-clones were hybridized for three days on slides pre-exposed to UV light after H-33258 staining. The following loci were mapped: GPR103 (BBU7q13, OAR6q13), TRAM1L1(OAR6q13dist), PPP3CA (BBU7q21, OAR6q15), SNCA (OAR6q17), PPARGC1A(BBU7q23, OAR6q17), UGDH (BBU7q25prox, OAR6q22prox), KDR (BBU7q27, OAR6q22), CNOT6L (BBU7q32prox, OAR6q32prox), NUP54 (BBU7q32, BBU6q32), DMP1 (BBU7q34dist-q36prox, OAR6q34dist-q36prox), QDPR (BBU7q36, OAR6q36). All loci mapped in homoeologous chromosomes and chromosome bands of the two species and their locations are in agreement with the previous RH-mapping performed on BBU7 with some difference in the distal region of BBU7. However, the present cytogenetic map better anchors the RH-map on specific river buffalo chromosome bands. In addition, eleven loci were assigned for the first time in sheep to OAR6, noticeably extending the cytogenetic map on this important chromosome which encodes caseins. Two loci (TRAM1L1 and SNCA) mapped in sheep were unmapped in river buffalo in three different FISH experiments. Comparisons between integrated cytogenetic maps of BBU7/OAR6 (and BTA6) with human chromosome 4 (HSA4) revealed complex chromosome rearrangements differentiating these chromosomes.