Refining the Camelus dromedarius Myostatin Gene Polymorphism through Worldwide Whole-Genome Sequencing.

Myostatin (MSTN) is a highly conserved negative regulator of skeletal muscle in mammals. Inactivating mutations results in a hyper-muscularity phenotype known as "double muscling" in several livestock and model species. In Camelus dromedarius, the gene structure organization and the sequence polymorphisms have been previously investigated, using Sanger and Next-Generation Sequencing technologies on a limited number of animals. Here, we carried out a follow-up study with the aim to further expand our knowledge about the sequence polymorphisms at the myostatin locus, through the whole-genome sequencing data of 183 samples representative of the geographical distribution range for this species. We focused our polymorphism analysis on the ±5 kb upstream and downstream region of the MSTN gene. A total of 99 variants (77 Single Nucleotide Polymorphisms and 22 indels) were observed. These were mainly located in intergenic and intronic regions, with only six synonymous Single Nucleotide Polymorphisms in exons. A sequence comparative analysis among the three species within the Camelus genus confirmed the expected higher genetic distance of C. dromedarius from the wild and domestic two-humped camels compared to the genetic distance between C. bactrianus and C. ferus. In silico functional prediction highlighted: (i) 213 differential putative transcription factor-binding sites, out of which 41 relative to transcription factors, with known literature evidence supporting their involvement in muscle metabolism and/or muscle development; and (ii) a number of variants potentially disrupting the canonical MSTN splicing elements, out of which two are discussed here for their potential ability to generate a prematurely truncated (inactive) form of the protein. The distribution of the considered variants in the studied cohort is discussed in light of the peculiar evolutionary history of this species and the hypothesis that extremely high muscularity, associated with a homozygous condition for mutated (inactivating) alleles at the myostatin locus, may represent, in arid desert conditions, a clear metabolic disadvantage, emphasizing the thermoregulatory and water availability challenges typical of these habitats.

Whole-genome analysis of introgressive hybridization and characterization of the bovine legacy of Mongolian yaks.

The yak is remarkable for its adaptation to high altitude and occupies a central place in the economies of the mountainous regions of Asia. At lower elevations, it is common to hybridize yaks with cattle to combine the yak's hardiness with the productivity of cattle. Hybrid males are sterile, however, preventing the establishment of stable hybrid populations, but not a limited introgression after backcrossing several generations of female hybrids to male yaks. Here we inferred bovine haplotypes in the genomes of 76 Mongolian yaks using high-density SNP genotyping and whole-genome sequencing. These yaks inherited ∼1.3% of their genome from bovine ancestors after nearly continuous admixture over at least the last 1,500 years. The introgressed regions are enriched in genes involved in nervous system development and function, and particularly in glutamate metabolism and neurotransmission. We also identified a novel mutation associated with a polled (hornless) phenotype originating from Mongolian Turano cattle. Our results suggest that introgressive hybridization contributed to the improvement of yak management and breeding.

WIDDE: a Web-Interfaced next generation database for genetic diversity exploration, with a first application in cattle.

BACKGROUND: The advent and democratization of next generation sequencing and genotyping technologies lead to a huge amount of data for the characterization of population genetic diversity in model and non model-species. However, efficient storage, management, cross-analyzing and exploration of such dense genotyping datasets remain challenging. This is particularly true for the bovine species where many SNP datasets have been generated in various cattle populations with different genotyping tools. DESCRIPTION: We developed WIDDE, a Web-Interfaced Next Generation Database that stands as a generic tool applicable to a wide range of species and marker types ( http://widde.toulouse.inra.fr). As a first illustration, we hereby describe its first version dedicated to cattle biodiversity, which includes a large and evolving cattle genotyping dataset for over 750,000 SNPs available on 129 (89 public) different cattle populations representative of the world-wide bovine genetic diversity and on 7 outgroup bovid species. This version proposes an optional marker and individual filtering step, an export of genotyping data in different popular formats, and an exploration of genetic diversity through a principal component analysis. Users can also explore their own genotyping data together with data from WIDDE, assign their samples to WIDDE populations based on distance assignment method and supervised clustering, and estimate their ancestry composition relative to the populations represented in the database. CONCLUSION: The cattle version of WIDDE represents to our knowledge the first database dedicated to cattle biodiversity and SNP genotyping data that will be very useful for researchers interested in this field. As a generic tool applicable to a wide range of marker types, WIDDE is overall intended to the genetic diversity exploration of any species and will be extended to other species shortly. The structure makes it easy to include additional output formats and new tools dedicated to genetic diversity exploration.

C-Nap1 mutation affects centriole cohesion and is associated with a Seckel-like syndrome in cattle.

Caprine-like Generalized Hypoplasia Syndrome (SHGC) is an autosomal-recessive disorder in Montbéliarde cattle. Affected animals present a wide range of clinical features that include the following: delayed development with low birth weight, hind limb muscular hypoplasia, caprine-like thin head and partial coat depigmentation. Here we show that SHGC is caused by a truncating mutation in the CEP250 gene that encodes the centrosomal protein C-Nap1. This mutation results in centrosome splitting, which neither affects centriole ultrastructure and duplication in dividing cells nor centriole function in cilium assembly and mitotic spindle organization. Loss of C-Nap1-mediated centriole cohesion leads to an altered cell migration phenotype. This discovery extends the range of loci that constitute the spectrum of autosomal primary recessive microcephaly (MCPH) and Seckel-like syndromes.

Whole-genome sequencing of 234 bulls facilitates mapping of monogenic and complex traits in cattle.

The 1000 bull genomes project supports the goal of accelerating the rates of genetic gain in domestic cattle while at the same time considering animal health and welfare by providing the annotated sequence variants and genotypes of key ancestor bulls. In the first phase of the 1000 bull genomes project, we sequenced the whole genomes of 234 cattle to an average of 8.3-fold coverage. This sequencing includes data for 129 individuals from the global Holstein-Friesian population, 43 individuals from the Fleckvieh breed and 15 individuals from the Jersey breed. We identified a total of 28.3 million variants, with an average of 1.44 heterozygous sites per kilobase for each individual. We demonstrate the use of this database in identifying a recessive mutation underlying embryonic death and a dominant mutation underlying lethal chrondrodysplasia. We also performed genome-wide association studies for milk production and curly coat, using imputed sequence variants, and identified variants associated with these traits in cattle.

Design and characterization of a 52K SNP chip for goats.

The success of Genome Wide Association Studies in the discovery of sequence variation linked to complex traits in humans has increased interest in high throughput SNP genotyping assays in livestock species. Primary goals are QTL detection and genomic selection. The purpose here was design of a 50-60,000 SNP chip for goats. The success of a moderate density SNP assay depends on reliable bioinformatic SNP detection procedures, the technological success rate of the SNP design, even spacing of SNPs on the genome and selection of Minor Allele Frequencies (MAF) suitable to use in diverse breeds. Through the federation of three SNP discovery projects consolidated as the International Goat Genome Consortium, we have identified approximately twelve million high quality SNP variants in the goat genome stored in a database together with their biological and technical characteristics. These SNPs were identified within and between six breeds (meat, milk and mixed): Alpine, Boer, Creole, Katjang, Saanen and Savanna, comprising a total of 97 animals. Whole genome and Reduced Representation Library sequences were aligned on >10 kb scaffolds of the de novo goat genome assembly. The 60,000 selected SNPs, evenly spaced on the goat genome, were submitted for oligo manufacturing (Illumina, Inc) and published in dbSNP along with flanking sequences and map position on goat assemblies (i.e. scaffolds and pseudo-chromosomes), sheep genome V2 and cattle UMD3.1 assembly. Ten breeds were then used to validate the SNP content and 52,295 loci could be successfully genotyped and used to generate a final cluster file. The combined strategy of using mainly whole genome Next Generation Sequencing and mapping on a contig genome assembly, complemented with Illumina design tools proved to be efficient in producing this GoatSNP50 chip. Advances in use of molecular markers are expected to accelerate goat genomic studies in coming years.

Novel insights into the bovine polled phenotype and horn ontogenesis in Bovidae.

Despite massive research efforts, the molecular etiology of bovine polledness and the developmental pathways involved in horn ontogenesis are still poorly understood. In a recent article, we provided evidence for the existence of at least two different alleles at the Polled locus and identified candidate mutations for each of them. None of these mutations was located in known coding or regulatory regions, thus adding to the complexity of understanding the molecular basis of polledness. We confirm previous results here and exhaustively identify the causative mutation for the Celtic allele (PC) and four candidate mutations for the Friesian allele (PF). We describe a previously unreported eyelash-and-eyelid phenotype associated with regular polledness, and present unique histological and gene expression data on bovine horn bud differentiation in fetuses affected by three different horn defect syndromes, as well as in wild-type controls. We propose the ectopic expression of a lincRNA in PC/p horn buds as a probable cause of horn bud agenesis. In addition, we provide evidence for an involvement of OLIG2, FOXL2 and RXFP2 in horn bud differentiation, and draw a first link between bovine, ovine and caprine Polled loci. Our results represent a first and important step in understanding the genetic pathways and key process involved in horn bud differentiation in Bovidae.

Fine mapping of copy number variations on two cattle genome assemblies using high density SNP array.

BACKGROUND: Btau_4.0 and UMD3.1 are two distinct cattle reference genome assemblies. In our previous study using the low density BovineSNP50 array, we reported a copy number variation (CNV) analysis on Btau_4.0 with 521 animals of 21 cattle breeds, yielding 682 CNV regions with a total length of 139.8 megabases. RESULTS: In this study using the high density BovineHD SNP array, we performed high resolution CNV analyses on both Btau_4.0 and UMD3.1 with 674 animals of 27 cattle breeds. We first compared CNV results derived from these two different SNP array platforms on Btau_4.0. With two thirds of the animals shared between studies, on Btau_4.0 we identified 3,346 candidate CNV regions representing 142.7 megabases (~4.70%) of the genome. With a similar total length but 5 times more event counts, the average CNVR length of current Btau_4.0 dataset is significantly shorter than the previous one (42.7 kb vs. 205 kb). Although subsets of these two results overlapped, 64% (91.6 megabases) of current dataset was not present in the previous study. We also performed similar analyses on UMD3.1 using these BovineHD SNP array results. Approximately 50% more and 20% longer CNVs were called on UMD3.1 as compared to those on Btau_4.0. However, a comparable result of CNVRs (3,438 regions with a total length 146.9 megabases) was obtained. We suspect that these results are due to the UMD3.1 assembly's efforts of placing unplaced contigs and removing unmerged alleles. Selected CNVs were further experimentally validated, achieving a 73% PCR validation rate, which is considerably higher than the previous validation rate. About 20-45% of CNV regions overlapped with cattle RefSeq genes and Ensembl genes. Panther and IPA analyses indicated that these genes provide a wide spectrum of biological processes involving immune system, lipid metabolism, cell, organism and system development. CONCLUSION: We present a comprehensive result of cattle CNVs at a higher resolution and sensitivity. We identified over 3,000 candidate CNV regions on both Btau_4.0 and UMD3.1, further compared current datasets with previous results, and examined the impacts of genome assemblies on CNV calling.

Design of a bovine low-density SNP array optimized for imputation.

The Illumina BovineLD BeadChip was designed to support imputation to higher density genotypes in dairy and beef breeds by including single-nucleotide polymorphisms (SNPs) that had a high minor allele frequency as well as uniform spacing across the genome except at the ends of the chromosome where densities were increased. The chip also includes SNPs on the Y chromosome and mitochondrial DNA loci that are useful for determining subspecies classification and certain paternal and maternal breed lineages. The total number of SNPs was 6,909. Accuracy of imputation to Illumina BovineSNP50 genotypes using the BovineLD chip was over 97% for most dairy and beef populations. The BovineLD imputations were about 3 percentage points more accurate than those from the Illumina GoldenGate Bovine3K BeadChip across multiple populations. The improvement was greatest when neither parent was genotyped. The minor allele frequencies were similar across taurine beef and dairy breeds as was the proportion of SNPs that were polymorphic. The new BovineLD chip should facilitate low-cost genomic selection in taurine beef and dairy cattle.

A newly described bovine type 2 scurs syndrome segregates with a frame-shift mutation in TWIST1.

The developmental pathways involved in horn development are complex and still poorly understood. Here we report the description of a new dominant inherited syndrome in the bovine Charolais breed that we have named type 2 scurs. Clinical examination revealed that, despite a strong phenotypic variability, all affected individuals show both horn abnormalities similar to classical scurs phenotype and skull interfrontal suture synostosis. Based on a genome-wide linkage analysis using Illumina BovineSNP50 BeadChip genotyping data from 57 half-sib and full-sib progeny, this locus was mapped to a 1.7 Mb interval on bovine chromosome 4. Within this region, the TWIST1 gene encoding a transcription factor was considered as a strong candidate gene since its haploinsufficiency is responsible for the human Saethre-Chotzen syndrome, characterized by skull coronal suture synostosis. Sequencing of the TWIST1 gene identified a c.148_157dup (p.A56RfsX87) frame-shift mutation predicted to completely inactivate this gene. Genotyping 17 scurred and 20 horned founders of our pedigree as well as 48 unrelated horned controls revealed a perfect association between this mutation and the type 2 scurs phenotype. Subsequent genotyping of 32 individuals born from heterozygous parents showed that homozygous mutated progeny are completely absent, which is consistent with the embryonic lethality reported in Drosophila and mouse suffering from TWIST1 complete insufficiency. Finally, data from previous studies on model species and a fine description of type 2 scurs symptoms allowed us to propose different mechanisms to explain the features of this syndrome. In conclusion, this first report on the identification of a potential causal mutation affecting horn development in cattle offers a unique opportunity to better understand horn ontogenesis.

The scurs inheritance: new insights from the French Charolais breed.

BACKGROUND: Polled animals are valued in cattle industry because the absence of horns has a significant economic impact. However, some cattle are neither polled nor horned but have so-called scurs on their heads, which are corneous growths loosely attached to the skull. A better understanding of the genetic determinism of the scurs phenotype would help to fine map the polled locus. To date, only one study has attempted to map the scurs locus in cattle. Here, we have investigated the inheritance of the scurs phenotype in the French Charolais breed and examined whether the previously proposed localisation of the scurs locus on bovine chromosome 19 could be confirmed or not. RESULTS: Our results indicate that the inheritance pattern of the scurs phenotype in the French Charolais breed is autosomal recessive with complete penetrance in both sexes, which is different from what is reported for other breeds. The frequency of the scurs allele (Sc) reaches 69.9% in the French Charolais population. Eleven microsatellite markers on bovine chromosome 19 were genotyped in 267 offspring (33 half-sib and full-sib families). Both non-parametric and parametric linkage analyses suggest that in the French Charolais population the scurs locus may not map to the previously identified region. A new analysis of an Angus-Hereford and Hereford-Hereford pedigree published in 1978 enabled us to calculate the frequency of the Sc allele in the Hereford breed (89.4%) and to study the penetrance of this allele in males heterozygous for both polled and scurs loci (40%). This led us to revise the inheritance pattern of the scurs phenotype proposed for the Hereford breed and to suggest that allele Sc is not fully but partially dominant in double heterozygous males while it is always recessive in females. Crossbreeding involving the Charolais breed and other breeds gave results similar to those reported in the Hereford breed. CONCLUSION: Our results suggest the existence of unknown genetics factors modifying the expression of the scurs locus in double heterozygous Hereford and Angus males. The specific inheritance pattern of the scurs locus in the French Charolais breed represents an opportunity to map this gene and to identify the molecular mechanisms regulating the growth of horns in cattle.

Chromosomal homology between the human and the bovine DMRT1 genes.

Doublesex and mab-3 related transcription factor 1 (DMRT1) is considered to be the most conserved gene among loci involved in the molecular pathways of animal sexual development. In the majority of the extensively examined vertebrates, its function is limited to the upstream or downstream testis regulators acting during embryogenesis. Our present study demonstrated the structural homology between DMRT1 orthologos in human and cattle. A BAC clone with a specific bovine sequence of the gene was used in the FISH mapping experiments. The physical localization of DMRT1 in cattle (BTA 8q17) was determined and its homology to the human locus was shown (HSA 9p24.3). Furthermore, another BAC probe, containing the sequence of the human homologue (pBACe3.6), generated hybridisation signals on bovine metaphase chromosomes and indicated the physical location of the autosomal bovine DMRT1 locus. Further investigations of the gene in domestic animals might provide more support for its conservative status and may help in understanding the molecular mechanisms involved in the occurrence of sexual abnormalities often diagnosed in livestock.

The bovine dilated cardiomyopathy locus maps to a 1.0-Mb interval on chromosome 18.

Cardiomyopathies are myocardial diseases that lead to cardiac dysfunction, heart failure, arrhythmia, and sudden death. In human medicine, cardiomyopathies frequently warrant heart transplantation in children and adults. Bovine dilated cardiomyopathy (BDCMP) is a heart muscle disorder that has been observed during the last 30 years in cattle of Holstein-Friesian origin. In Switzerland BDCMP affects Swiss Fleckvieh and Red Holstein breeds. BDCMP is characterized by a cardiac enlargement with ventricular remodeling and chamber dilatation. The common symptoms in affected animals are subacute subcutaneous edema, congestion of the jugular veins, and tachycardia with gallop rhythm. A cardiomegaly with dilatation and hypertrophy of all heart chambers, myocardial degeneration, and fibrosis are typical postmortem findings. It was shown that all BDCMP cases reported worldwide traced back to a red factor-carrying Holstein-Friesian bull, ABC Reflection Sovereign. An autosomal recessive mode of inheritance was proposed for BDCMP. Recently, the disease locus was mapped to a 6.7-Mb interval MSBDCMP06-BMS2785 on bovine Chr 18 (BTA18). In the present study the BDCMP locus was fine mapped by using a combined strategy of homozygosity mapping and association study. A BAC contig of 2.9 Mb encompassing the crucial interval was constructed to establish the correct marker order on BTA18. We show that the disease locus is located in a gene-rich interval of 1.0 Mb and is flanked by the microsatellite markers DIK3006 and MSBDCMP51.

Fine mapping of quantitative trait loci affecting female fertility in dairy cattle on BTA03 using a dense single-nucleotide polymorphism map.

Fertility quantitative trait loci (QTL) are of high interest in dairy cattle since insemination failure has dramatically increased in some breeds such as Holstein. High-throughput SNP analysis and SNP microarrays give the opportunity to genotype many animals for hundreds SNPs per chromosome. In this study, due to these techniques a dense SNP marker map was used to fine map a QTL underlying nonreturn rate measured 90 days after artificial insemination previously detected with a low-density microsatellite marker map. A granddaughter design with 17 Holstein half-sib families (926 offspring) was genotyped for a set of 437 SNPs mapping to BTA3. Linkage analysis was performed by both regression and variance components analysis. An additional analysis combining both linkage analysis and linkage-disequilibrium information was applied. This method first estimated identity-by-descent probabilities among base haplotypes. These probabilities were then used to group the base haplotypes in different clusters. A QTL explaining 14% of the genetic variance was found with high significance (P < 0.001) at position 19 cM with the linkage analysis and four sires were estimated to be heterozygous (P < 0.05). Addition of linkage-disequilibrium information refined the QTL position to a set of narrow peaks. The use of the haplotypes of heterozygous sires offered the possibility to give confidence in some peaks while others could be discarded. Two peaks with high likelihood-ratio test values in the region of which heterozygous sires shared a common haplotype appeared particularly interesting. Despite the fact that the analysis did not fine map the QTL in a unique narrow region, the method proved to be able to handle efficiently and automatically a large amount of information and to refine the QTL position to a small set of narrow intervals. In addition, the QTL identified was confirmed to have a large effect (explaining 13.8% of the genetic variance) on dairy cow fertility as estimated by nonreturn rate at 90 days.

A physical map of the bovine genome.

BACKGROUND: Cattle are important agriculturally and relevant as a model organism. Previously described genetic and radiation hybrid (RH) maps of the bovine genome have been used to identify genomic regions and genes affecting specific traits. Application of these maps to identify influential genetic polymorphisms will be enhanced by integration with each other and with bacterial artificial chromosome (BAC) libraries. The BAC libraries and clone maps are essential for the hybrid clone-by-clone/whole-genome shotgun sequencing approach taken by the bovine genome sequencing project. RESULTS: A bovine BAC map was constructed with HindIII restriction digest fragments of 290,797 BAC clones from animals of three different breeds. Comparative mapping of 422,522 BAC end sequences assisted with BAC map ordering and assembly. Genotypes and pedigree from two genetic maps and marker scores from three whole-genome RH panels were consolidated on a 17,254-marker composite map. Sequence similarity allowed integrating the BAC and composite maps with the bovine draft assembly (Btau3.1), establishing a comprehensive resource describing the bovine genome. Agreement between the marker and BAC maps and the draft assembly is high, although discrepancies exist. The composite and BAC maps are more similar than either is to the draft assembly. CONCLUSION: Further refinement of the maps and greater integration into the genome assembly process may contribute to a high quality assembly. The maps provide resources to associate phenotypic variation with underlying genomic variation, and are crucial resources for understanding the biology underpinning this important ruminant species so closely associated with humans.

Genetic and haplotypic structure in 14 European and African cattle breeds.

To evaluate and compare the extent of LD in cattle, 1536 SNPs, mostly localized on BTA03, were detected in silico from available sequence data using two different methods and genotyped on samples from 14 distinct breeds originating from Europe and Africa. Only 696 SNPs could be validated, confirming the importance of trace-quality information for the in silico detection. Most of the validated SNPs were informative in several breeds and were used for a detailed description of their genetic structure and relationships. Results obtained were in agreement with previous studies performed on microsatellite markers and using larger samples. In addition, the majority of the validated SNPs could be mapped precisely, reaching an average density of one marker every 311 kb. This allowed us to analyze the extent of LD in the different breeds. Decrease of LD with physical distance across breeds revealed footprints of ancestral LD at short distances (<10 kb). As suggested by the haplotype block structure, these ancestral blocks are organized, within a breed, into larger blocks of a few hundred kilobases. In practice, such a structure similar to that already reported in dogs makes it possible to develop a chip of <300,000 SNPs, which should be efficient for mapping purposes in most cattle breeds.

Position independent and copy-number-related expression of the bovine neonatal Fc receptor alpha-chain in transgenic mice carrying a 102 kb BAC genomic fragment.

We generated and characterized transgenic mice carrying a 102 kb bovine genomic fragment, encoding the neonatal Fc receptor alpha-chain (bFcRn). FcRn plays a crucial role in the maternal IgG transport and it also regulates the IgG and albumin homeostasis. Some of its functions and transcriptional regulation show species specific differences. The FcRn heterodimer is composed of the alpha-chain and beta-2-microglobulin (beta2 m). A bacterial artificial chromosome containing the bovine FcRn alpha-chain gene (bFCGRT) with its 44 kb 5' and 50 kb long 3' flanking sequences was microinjected into fertilized mouse oocytes. Two of the transgenic lines generated, showed copy number related and integration site independent bFcRn expression. The bFcRn alpha-chain forms a functional receptor with the mouse beta2-microglobulin and extends the half-life of the mouse IgG in transgenic mice. Our results underline the feasibility of creating BAC transgenic mouse models of economically important bovine genes.

Congenital syndactyly in cattle: four novel mutations in the low density lipoprotein receptor-related protein 4 gene (LRP4).

BACKGROUND: Isolated syndactyly in cattle, also known as mulefoot, is inherited as an autosomal recessive trait with variable penetrance in different cattle breeds. Recently, two independent mutations in the bovine LRP4 gene have been reported as the primary cause of syndactyly in the Holstein and Angus cattle breeds. RESULTS: We confirmed the previously described LRP4 exon 33 two nucleotide substitution in most of the affected Holstein calves and revealed additional evidence for allelic heterogeneity by the identification of four new LRP4 non-synonymous point mutations co-segregating in Holstein, German Simmental and Simmental-Charolais families. CONCLUSION: We confirmed a significant role of LRP4 mutations in the pathogenesis of congenital syndactyly in cattle. The newly detected missense mutations in the LRP4 gene represent independent mutations affecting different conserved protein domains. However, the four newly described LRP4 mutations do still not explain all analyzed cases of syndactyly.

Cloning of the bovine prion-like Shadoo (SPRN) gene by comparative analysis of the predicted genomic locus.

SPRN is a new prion-like gene coding for Sho, a protein with significant similarity to PrP. SPRN was initially described in zebrafish; however, the strong evolutionary conservation led to the hypothesis that SPRN might be the ancestral prion-like gene. We mapped SPRN in Bos taurus by comparative analysis of the locus and of the predicted flanking genes. BACs, spanning the whole SPRN genomic locus, were assigned to BTA26q23 by radiation hybrid mapping and fluorescent in situ hybridization (FISH). Sequencing of five genes flanking SPRN, namely, ECHS1, PAOX, MTG1, SPRN, and CYP2E1, high-resolution FISH on mechanically stretched chromosomes, and combed BAC DNA allowed us to establish their order and reciprocal orientation. The results confirmed that BTA26q23 corresponds to HSA10q24.3-26.3, which is the site where the human SPRN is located. The gene order in Bos taurus is the same as in man, cen-ECHS1-PAOX-MTG1-SPRN-CYP2E1-tel, but PAOX has a different orientation in the two species. SPRN has the typical two-exon PRNP arrangement, with the CDS fully contained within exon 2; furthermore, it codes for a 143-amino-acid protein with 74.8% identity and 84.7% similarity with the human PRNP. RT-PCR and Northern blot analysis showed that SPRN is expressed at high levels in brain and less in testis and lung.