Synteny mapping in the bovine: genes from human chromosome 4.

Genes homologous to those located on human chromosome 4 (HSA4) were mapped in the bovine to determine regions of syntenic conservation among humans, mice, and cattle. Previous studies have shown that two homologs of genes on HSA4, PGM2 and PEPS, are located in bovine syntenic group U15 (chromosome 6). The homologous mouse genes, Pgm-1 and Pep-7, are on MMU5. Using a panel of bovine x hamster hybrid somatic cells, we have assigned homologs of 11 additional HSA4 loci to their respective bovine syntenic groups. D4S43, D4S10, QDPR, IGJ, ADH2, KIT, and IF were assigned to syntenic group U15. This syntenic arrangement is not conserved in the mouse, where D4s43, D4s10, Qdpr, and Igj are on MMU5 while Adh-2 is on MMU3. IL-2, FGB, FGG, and F11, which also reside on MMU3, were assigned to bovine syntenic group U23. These data suggest that breaks and/or fusions of ancestral chromosomes carrying these genes occurred at different places during the evolution of humans, cattle, and mice.     

Synteny mapping in the bovine: genes from human chromosome 5.

In an effort to generate a more complete bovine syntenic map of Type I comparative anchor loci, seven homologs to genes found on HSA5 were mapped using a panel of bovine x rodent hybrid somatic cells. Five HSA5 genes, CSF2, RPS14, PDGFRB, FGFA, and CSF1R, were assigned to bovine syntenic group U22 (chromosome 7), while two others, C9 and HGMCR, mapped to U10 and U5, respectively. Previous studies had assigned the HSA5 marker SPARC to bovine syntenic group U22. The mapping of genes spanning the length of HSA5 in cattle and also in mouse permits syntenic comparisons between prototypic genomes of three mammalian orders, providing insight into the evolutionary history of this region of the ancestral mammalian genome.     

Mapping of bovine prolactin and rhodopsin genes in hybrid somatic cells

The genes encoding bovine prolactin and rhodopsin were assigned to syntenic groups on the basis of hybridization of DNA from a panel of bovine-hamster hybrid somatic cell lines with cloned prolactin and rhodopsin gene probes. Prolactin was found to be syntenic with previously mapped glyoxalase, BoLA and 21-hydroxylase genes, establishing a syntenic conservation with human chromosome 6. The presence of bovine rhodopsin sequences among the various hybrid cell lines was not concordant with any gene previously assigned to one of the 23 defined autosomal syntenic groups. Thus, rhodopsin marks a new bovine syntenic group, U24, leaving only five cattle autosomes unmarked by at least one biochemical or molecular marker.     

Chromosomal localization and detection of DNA polymorphisms in the bovine polymeric immunoglobulin receptor gene

Polymeric immunoglobulin receptor (PIGR) mediates transcellular transport of secretory antibodies in glandular and mucosal epithelial cells. By use of a bovine-rodent somatic cell hybrid panel the bovine PIGR locus has been assigned to syntenic group U1. Using in situ hybridization, PIGR was localized to bovine chromosome 16, segment q13, thus confirming the recent assignment of syntenic group U1 to this chromosome. Two common restriction fragment length polymorphisms (RFLPs) with the enzymes Bam HI and MspI were detected using the PIGR cDNA as probe. Direct PCR sequencing of a segment in the PIGR coding region (nucleotides 162–413) from 13 bulls of Norwegian Cattle revealed single nucleotide exchanges at two positions. An efficient PCR-RFLP method for detection of these mutations was developed.     

Cytogenetical anchoring of sheep linkage map and syntenic groups using a sheep BAC library

In order to simultaneously integrate linkage and syntenic groups to the ovine chromosomal map, a sheep bacterial artificial chromosome (BAC) library was screened with previously assigned microsatellites using a sheep-hamster hybrid panel and genetic linkage. Thirty-three BACs were obtained, fluorescently labelled and hybridised on sheep-goat hybrid metaphases (2n = 57). This study allowed us, (i), to anchor all linkage groups on sheep chromosomes, (ii), to give information on the probable position of the centromere on the linkage map for the centromeric chromosomes, (iii), to contradict the previous orientation of the ovine × linkage group by the mapping of BMS1008 on OARXq38. Concerning our somatic cell hybrid panel, this study resulted in the assignment of all the previously unassigned groups to ovine chromosomes and a complete characterisation of the hybrid panel. In addition, since hybridisations were performed on a sheep-goat hybrid, new marker/anchoring points were added to the caprine cytogenetic map.     

The thyroglobulin gene is syntenic with the MYC and MOS protooncogenes and carbonic anhydrase II and maps to chromosome 14 in cattle

Using a panel of bovine x Chinese hamster hybrid somatic cells, sequences homologous to genes spanning human chromosome arm 8q have been syntenically assigned in cattle. Thyroglobulin (TG), carbonic anhydrase II (CA2), and the protooncogenes MYC and MOS were assigned to a newly identified bovine syntenic group, U23. Additionally, in situ hybridization of the thyroglobulin probe to bovine metaphase chromosomes revealed this syntenic group to be on bovine chromosome 14 and the bovine thyroglobulin gene to reside at 14q12----q15.     

Physical and linkage mapping of the bovine genome with cosmids

We have initiated a mapping strategy using cosmid clones to chromosomally anchor a high-resolution bovine genetic linkage map. Ten cosmids containing microsatellites were assigned to bovine chromosomes by fluorescence in situ suppression hybridization (FISH). Four cosmid clones, three of which contain an informative microsatellite, were assigned to autosomes 5, 13, 24, and 28. The assignment to autosome 13 anchors bovine syntenic group U11. Two additional cosmid clones, each containing informative microsatellites, are assigned to autosomes 9 and 29, auchoring bovine linkage groups U2 and U8, respectively. Four cosmid clones, three of which contain informative microsatellites, also provide the first assignment to autosome 25, anchoring bovine syntenic group U7 and orienting the corresponding linkage group relative to the centromere.     

Mapping HSA 3 loci in cattle: additional support for the ancestral synteny of HSA 3 and 21.

Homologs to genes residing on human chromosome 3 (HSA 3) map to four mouse chromosomes (MMU) 3, 6, 9, and 16. In the bovine, two syntenic groups that contain HSA 3 homologs, unassigned syntenic groups 10 (U10) and 12 (U12), have been defined. U10 also contains HSA 21 genes, which is similar to the situation seen on MMU 16, whereas U12 apparently contains only HSA 3 homologs. The syntenic arrangement of other HSA 3 homologs in the bovine was investigated by physically mapping five genes through segregation analysis of a bovine-hamster hybrid somatic cell panel. The genes mapped include Friend-murine leukemia virus integration site 3 homolog (FIM3; HSA 3/MMU 3), sucrase-isomaltase (SI) and glutathione peroxidase 1 (GPX1) (HSA 3/MMU ?), murine leukemia viral (v-raf-1) oncogene homolog 1 (RAF1; HSA 3/MMU 6), and ceruloplasmin (CP; HSA 3/MMU 9). FIM3, SI, and CP mapped to bovine syntenic group U10, while RAF1 and GPX1 mapped to U12.     

Comparative mapping of twenty-eight bovine loci in sheep (Ovis aries, 2n = 54) and river buffalo (Bubalus bubalis, 2n = 50) by FISH

Peripheral blood cell cultures were treated for late incorporation of both BrdU and Hoechst-33258 to obtain R-banding pattern preparations. Twenty-eight bovine cosmids from 19 bovine syntenic groups (U), three of which contain type I loci and 25 which contain microsatellite loci and have previously been assigned to cattle chromosomes, were comparatively FISH-mapped to sheep and river buffalo chromosomes according to the standard karyotypes (13 loci for the first time in the latter species). The results enrich the physical maps of both species with information relative to the following loci and to the corresponding syntenic groups: IDVGA35 and IDVGA53 (U6), IDVGA61 and IDVGA84 (U13), JAB10 (U5), IDVGA41 and IDVGA57 (U27), IDVGA87 (U11), IDVGA32 and IDVGA10 (U19), IDVGA49, IDVGA66 and IDVGA68 (U1), ZNF164 (U23), IDVGA74 and IDVGA70 (U9), IDVGA47, IDVGA46 and IDVGA58 (U21), MAP1B (U14), IDVGA79 (U4), CATHL (U12), IDVGA71 (U8), IDVGA59 (U26), IDVGA29 (U29), IDVGA7 (U7), IDVGA82 (X), IDVGA50 (Y). All mapped loci were localized on homoeologous chromosomes and chromosome regions of the two species, confirming the high degree of chromosome homoeologies between the subfamilies Bovinae and Caprinae.     

Physically mapped,cosmid-derived microsatellite markers as anchor loci on bovine chromosomes

To identify physical and genetic anchor loci on bovine chromosomes, 13 cosmids, obtained after the screening of partial bovine cosmid libraries with the (CA)_n microsatellite motif, were mapped by fluorescence in situ hybridization (FISH). Eleven cosmid probes yielded a specific signal on one of the bovine chromosomes and identified the following loci: D5S2, D5S3, D6S3, D8S1, D11S5, D13S1, D16S5, D17S2, D19S2, D19S3, D21S8. Two cosmids produced centromeric signals on many chromosomes. The microsatellite-containing regions were subcloned and sequenced. The sequence information revealed that the two centromeric cosmids were derived from bovine satellites 1.723 and 1.709, respectively. A cosmid located in the subtelomeric region of Chromosome (Chr) 17 (D17S2) had features of a chromosome-specific satellite. Primers were designed for eight of the nonsatellite cosmids, and seven of these microsatellites were polymorphic with between three and eight alleles on a set of outbred reference families. The polymorphic and chromosomally mapped loci can now be used to physically anchor other bovine polymorphic markers by linkage analysis. The microsatellite primers were also applied to DNA samples of a previously characterized panel of somatic hybrid cell lines, allowing the assignment of seven microsatellite loci to defined syntenic groups. These assignments confirmed earlier mapping results, revealed a probable case of false synteny, and placed two formerly unassigned syntenic groups on specific chromosomes.     

Fifteen new synteny assignments of microsatellites to the bovine genome

A panel of 36 hamster-bovine hybrid cell lines was used to assign 15 bovine microsatellites. Locus identification, synteny group and/or chromosome assignment and registration number were as follows: ILSTS001 (U22, Chr07, D7S13), ILSTS002 (U09, Chr18, D18S7), ILSTS004 (U10, Chr01, D1S28), ILSTS005 (U05, Chr10, D10S25). ILSTS006 (U22, Chr07, D7S8), ILSTS008 (U24, Chr14, D14S15), ILSTS010 (U27, DU27S11), ILSTS011 (U24, Chr14, D14S16), ILSTS012 (U16, Chr11, D11S26), ILSTS015 (U07, Chr25, D25S3), ILSTS016 (U04, Chr21, D21S22), ILSTS017 (X, DXS11), ILSTS018 (U15, Chr06, D6S15), ILSTS019 (U07, Chr25, D25S7) and ILSTS020 (U05, Chr10, D10S27). These results contribute to the international effort to improve the bovine genetic map.     

A synteny map of the horse genome comprised of 240 microsatellite and RAPD markers

To generate a domestic horse genome map we integrated synteny information for markers screened on a somatic cell hybrid (SCH) panel with published information for markers physically assigned to chromosomes. The mouse-horse SCH panel was established by fusing pSV2neo transformed primary horse fibroblasts to either RAG or LMTk mouse cells, followed by G418 antibiotic selection. For each of the 108 cell lines of the panel, we defined the presence or absence of 240 genetic markers by PCR, including 58 random amplified polymorphic DNA (RAPD) markers and 182 microsatellites. Thirty-three syntenic groups were defined, comprised of two to 26 markers with correlation coefficient (r) values ranging from 0.70 to 1.0. Based on significant correlation values with physically mapped microsatellite (type II) or gene (type I) markers, 22 syntenic groups were assigned to horse chromosomes (1, 2, 3, 4, 6, 9, 10, 11, 12, 13, 15, 18, 19, 20, 21, 22, 23, 24, 26, 30, X and Y). The other 11 syntenic groups were provisionally assigned to the remaining chromosomes based on information provided by heterologous species painting probes and work in progress with type I markers.     

Comparative mapping ofIGHG,IGHM, FES,andFOS in domestic cattle

The immunoglobulin genes have not been genetically characterized as thoroughly in cattle as in other mammals, particularly humans and mice. Comparative gene mapping in mammals suggests that the bovine immunoglobulin heavy chain genes,IGHG4 andIGHM might be syntenic with theFOS oncogene. Interestingly, however, when these genes were assigned to bovine syntenic groups utilizing a panel of bovine: hamster hybrid somatic cells,IGH genes were shown to be syntenic with theFES oncogene rather thanFOS. In this studyIGH andFES were assigned toBos taurus chromosome 21 whileFOS was assigned to chromosome 10. In addition, bovine-specific immunoglobulin-like sequences were observed in the hybrid somatic cells, and one, IGHML1, was mapped to bovine syntenic group U16. The probes used for somatic-cell mapping were also used to screen a small number of cattle of several different breeds for restriction fragment length polymorphisms.IGHG4 andIGHM were shown to be highly polymorphisms. whileFOS andFES were not. Address correspondence and offprint requests to: J. E. Womack.     

Physical mapping confirms that sheep chromosome 10 has extensive conserved synteny with cattle chromosome 12 and human chromosome 13

The following loci, on human chromosome 13, have been newly assigned to sheep chromosome 10 using chromosomally characterized sheep-hamster cell hybrids: gap junction protein, beta 2, 26 kDa (connexin 26) (GJB2); gap junction protein, alpha 3, 46 kDa (connexin 46) (GJA3), and esterase D/formylglutathione hydrolase (ESD). This assignment of ESD is consistent with comparative mapping evidence, but not with an earlier report of it on sheep chromosome 3p26-p24. Cell hybrid analysis confirmed the location of another human chromosome 13 locus, retinoblastoma 1 (including osteosar-coma) (RBI), and the anonymous ovine genomic sequence RP11 on sheep chromosome 10. Isotopic in situ hybridization was used to regionally localize RP11 on to sheep 10q15-q22. The location of microsatellites AGLA226, OarDB3, OarHH41, OarVH58, and TGLA441, previously assigned to sheep chromosome 10 by linkage analysis, was confirmed by polymerase chain reaction using the cell hybrid panel. These mapping data provide further evidence that sheep chromosome 10 is the equivalent of cattle chromosome 12, and that these chromosomes show extensive conserved synteny with human chromosome 13.     

Genomic analysis of the major bovine milk protein genes.

The genomic arrangement of the major bovine milk protein genes has been determined using a combination of physical mapping techniques. The major milk proteins consist of the four caseins, alpha s1 (CASAS1), alpha s2 (CASAS2), beta (CASB), and kappa (CASK), as well as the two major whey proteins, alpha-lactalbumin (LALBA) and beta-lactoglobulin (LGB). A panel of bovine X hamster hybrid somatic cells analyzed for the presence or absence of bovine specific restriction fragments revealed the genes coding for the major milk proteins to reside on three chromosomes. The four caseins were assigned to syntenic group U15 and localized to bovine chromosome 6 at q31-33 by in situ hybridization. LALBA segregated with syntenic group U3, while LGB segregated with U16. Pulsed-field gel electrophoresis confirmed genetic mapping results indicating tight linkage of the casein genes. The four genes reside on less than 200 kb of DNA in the order CASAS1-CASB-CASAS2-CASK. Multiple restriction fragment length polymorphisms were also found at the six loci in three breeds of cattle.     

Synteny mapping in river buffalo

The cosegregation of ten coding loci has been investigated, in a panel of 37 somatic cell hybrids resulting from the fusion of a hamster cell line and river buffalo lymphocytes, by use of Southern hybridization technique. Five syntenic groups, TCRB-PGY3, ASS-ABL, FUCA1P-CRYG, MBP-YES1, and CGN1-ACTA1, previously assigned to cattle as U13, U16, U17, U28, and U29 respectively, were also found to be syntenic in buffalo. Based on the extensive syntenic conservation and banding homology between cattle and river buffalo, comparative mapping predicts the localization of these syntenic groups on river buffalo Chromosomes (Chrs) :BBU7, BBU12, BBU2q, BBU22, and BBU4q respectively as they have been previously localized on cattle Chrs BTA4, BTA11, BTA2, BTA24 & BTA28. Received: 2 April 1996 / Accepted: 4 July 1996     

Physical mapping of the lysozyme gene family in cattle

Amplification of an ancestral lysozyme gene in artiodactyls is associated with the evolution of foregut fermetation in the ruminant lineage and has resulted in about ten lysozyme genes in true ruminants. Hybridization of a cow stomach lysozyme 2 cDNA clone to restricted DNAs of a panel of cowxhamster hybrid cell lines revealed that all but one of the multiple bovine-specific bands segregate concordantly with the marker for bovine syntenic group U3 [Chromosome (Chr) 5]. The anomalous band was subsequently mapped to bovine syntenic group U22 (Chr 7) with a second panel of hybrids representing all 31 bovine syntenic groups. By two-dimensional pulsed-field gel electrophoresis the lysozyme genes on cattle Chr 5 were shown to be clustered on a 2- to 3-Mb DNA fragment, while the lactalbumin gene and pseudogenes that are paralogous and syntenic with the lysozymes were outside the lysozyme gene cluster. Chromosomal fluorescence in situ hybridization of a cocktail of lysozyme genomic clones localized the lysozyme gene cluster to cattle Chr 5 band 23, corroborating the somatic cell assignment.     

Somatic cell mapping of T-cell receptor CD3 complex and CD8 genes in cattle

Bovine genes encoding T-cell receptor, CD3, and CD8 molecules have been mapped to syntetic groups using bovine × rodent hybrid somatic cells. T-cell receptor and chains were assigned to bovine syntenic group U5, and the and genes were syntenic with each other and with markers on U13. CD3E and CD3D genes were syntenic with each other and located to bovine syntenic group U19. CD8 was most concordant with markers of syntenic group U16, although the concordancy was only 85% and the assignment must be regarded as tentative. The comparative gene maps of human chromosome 7, bovine syntenic group U13, and mouse chromosomes 6 and 13 suggest extensive evolutionary conservation.     

Four new DNA markers are assigned to the WAGR region of 11p13: isolation and regional assignment of 112 chromosome 11 anonymous DNA segments

One hundred eighty-three human single copy clones were isolated from the Livermore Laboratory chromosome 11 library (ID code LL11NSO1) and 112 of them were mapped to chromosome 11. Using a panel of somatic cell hybrids segregating chromosome 11 translocations and short arm deletions, 54 of the clones were assigned to one of nine segments on the short arm of chromosome 11; the remainder were assigned to the long arm. Nine of these clones map to 11p13, and four of the nine [57(D11S89), 530(D11S90), 706(D11S93), and 1104(D11S95)] are confined to the same segment within p13 that contains catalase (CAT), the beta subunit of follicle stimulating hormone (FSHB), and the Wilms' tumor-aniridia (WAGR) gene complex.     

Somatic cell mapping of bovine EC-SOD and SOD1L loci.

cDNA probes of human extracellular superoxide dismutase (EC-SOD) and bovine superoxide dismutase 1 (SOD1) genes were hybridized to Southern blots containing genomic DNAs from cow-rodent somatic cell lines segregating bovine chromosomes. The SOD1 probe identified two loci: the coding locus (SOD1), which mapped to bovine U10; and a related locus (SOD1L), which mapped to U11. EC-SOD mapped to bovine U15. The mapping of EC-SOD to human chromosome 4, and our mapping of EC-SOD to U15, further defines a region of extensive syntenic conservation between humans and domestic cows.