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Topic:DNA
DNA in horses refers to the genetic material that carries the hereditary information necessary for the growth, development, functioning, and reproduction of equine species. It consists of sequences of nucleotides that encode the genetic instructions used in the development and functioning of horses. DNA analysis in horses can provide insights into genetic diversity, lineage, and breed characteristics. It is also utilized in identifying genetic disorders, understanding hereditary traits, and assisting in selective breeding programs. This page compiles peer-reviewed research studies and scholarly articles that explore the structure, function, and applications of DNA analysis in equine genetics and breeding.
Equine severe combined immunodeficiency: a defect in V(D)J recombination and DNA-dependent protein kinase activity. V(D)J rearrangement is the molecular mechanism by which an almost infinite array of specific immune receptors are generated. Defects in this process result in profound immunodeficiency as is the case in the C.B-17 SCID mouse or in RAG-1 (recombination-activating gene 1) or RAG-2 deficient mice. It has recently become clear that the V(D)J recombinase most likely consists of both lymphoid-specific factors and ubiquitously expressed components of the DNA double-strand break repair pathway. The deficit in SCID mice is in a factor that is required for both of these pathways. In this report, we show...
Cloning of a DNA repeat element from horse: DNA sequence and chromosomal localization. A DNA repeat element, revealed initially by digestion of horse DNA with TaqI, was cloned and characterized by Southern and in situ hybridization studies and nucleotide sequencing. The clone, e4/1, consisted of 32 tandem reiteration of a unit repeat of 21-22 bp, and produced multilocus DNA fingerprinting profiles that were useful for parentage analysis in horses. The tandem repeat element was shown by in situ hybridization to be localized in the centromeres of the acrocentric but not metacentric classes of horse chromosomes.
Identification of opossums (Didelphis virginiana) as the putative definitive host of Sarcocystis neurona. Sarcocystis neurona is an apicomplexan that causes equine protozoal myeloencephalitis (EPM) in North and South America. Horses appear to be an aberrant host, because the merozoites continually divide in the central nervous system, without encysting. The natural host species has not previously been identified. The small subunit ribosomal RNA (SSURNA) gene of S. neurona was compared to those of Sarcocystis muris, Sarcocystis cruzi, Toxoplasma gondii, and Cryptosporidium parvum to identify a unique region suitable for a species-specific amplification primer. The S. neurona SSURNA primer was used ...
Families of tandemly repeated DNA elements from horse: cloning, nucleotide sequence, and organization. DNA repeats, revealed initially by digestion of horse DNA with restriction enzymes, were cloned and characterized by cross-hybridization studies and nucleotide sequencing. The Sau-like family of tandem repeats contained two classes of repetitive elements with unit repeats of about 80 bp that shared no sequence similarity. Both unit repeats were present, frequently in tandem, in cloned segments of horse DNA of less than 600 bp. Evidence is presented, based on their ladderlike patterns of hybridization to horse DNA and their high level of similarity to published sequences of satellites from equi...
Sarcocystis falcatula from passerine and psittacine birds: synonymy with Sarcocystis neurona, agent of equine protozoal myeloencephalitis. Equine protozoal myeloencephalitis (EPM) is a neurologic disease of horses caused by Sarcocystis neurona. The horse is a dead-end host for S. neurona and the definitive and intermediate hosts have not previously been identified. We hypothesized that S. neurona is actually Sarcocystis falcatula, a parasite that cycles in nature between Virginia opossums (Didelphis virginiana) and any of a variety of avian intermediate hosts. We extracted DNA from S. falcatula sarcocysts in the muscle of a brown-headed cowbird (Molothrus ater) and from schizonts in a fixed specimen of lung from a Moluccan cockat...
Initiation of transcription and nucleologenesis in equine embryos. The time of activation of the embryonic genome (maternal-embryonic transition) in equine embryos was investigated by assessing incorporation of 3H-uridine and nucleolar development. In Experiment 1, embryos were recovered from the oviduct (n = 15) and the uterus (n = 3). Recovered embryos were assessed for morphologic development and quality score. Recovered embryos with less than 8 cells (two cells, n = 4; four cells, n = 5; five cells, n = 2) were incubated with 3H-uridine (560 microCi/ml) for 10 hr, while eight-cell embryos (n = 2), morulae (n = 2), and blastocysts (n = 3) were incubated wi...
Demonstration of tissue-specific promoters in nonprimate species that express aromatase P450 in placentae. Conversion of androgens to estrogens is catalyzed by aromatase P450 (P450arom; the product of the CYP19 gene). Regulation of tissue-specific expression of P450arom in humans is due, in part, to alternative transcriptional start sites that arise as a consequence of the use of granulosa cells and placental tissue from cows, horses, and pigs (ungulates) in order to determine whether these species, like the human, utilize tissue-specific promoters to drive P450arom expression. The majority of transcripts in the placenta have 5'-termini that differ from those in the ovary upstream of a common site ...
Molecular cloning of equine interleukin-1 alpha and -beta cDNAs. Equine interleukin-1 alpha (IL-1 alpha) and IL-1 beta were molecularly cloned to establish a basis for research on inflammatory and immune responses in the horse. Equine peripheral blood mononuclear cells (PBMC) were stimulated with lipopolysaccharide (LPS), and cDNA clones of equine IL-1 alpha and IL-1 beta covering the whole coding sequences were isolated from them. These equine IL-1 alpha and IL-1 beta clones contained open reading frames encoding 271 and 269 amino acids, respectively. The deduced amino acid sequence of equine IL-1 alpha showed 71.6% and 60.2% similarity with that of human ...
Sex diagnosis of equine preimplantation embryos using the polymerase chain reaction. A rapid and reliable method for sex determination of preimplantation-stage equine embryos has not been available. The aim of the present study was to find an enzyme which would distinguish sexes in the horse by finding a polymorphic restriction site between the ZFY and ZFX homologues amplified by the polymerase chain reaction (PCR). Altogether, 38 different restriction enzymes were tested using female and male DNA extracted from blood. The primers used for amplification were selected from conserved sequences between human ZFY and ZFX genes and mouse Zfy-1 and Zfy-2 genes. Nine enzymes cut the ...
Molecular cloning of DNA for inhibin alpha-subunit from equine ovary. cDNA encoding equine inhibin alpha-subunit precursor protein was isolated from an equine ovarian cDNA library. For screening, the DNA probe was amplified by the RT-PCR using primers designed based on the rat inhibin alpha-subunit cDNA sequence. Out of 1.2 x 10(5) plaques screened, 19 positive clones were isolated, and one of these clones (Eq-alpha-11) contained a complete open reading frame encoding 367 amino acids. The similarity of the deduced amino acid sequences of both equine inhibin alpha-subunit precursor protein and the mature protein were greater than 80% to those of other six mammali...
Partial sequence of the equine immunoglobulin epsilon heavy chain cDNA. In order to isolate a part of the immunoglobulin E (IgE) heavy chain cDNA of the horse, primers have been designed based upon well conserved sequences in humans, sheep and rats. The PCR resulted in a 500 bp fragment which hybridised with a human IgE constant region probe. The fragment was cloned and sequenced and its derived protein sequence compared with the corresponding sequences in humans, sheep and mice. Most amino acids common to these three species are also shared by the horse.
Mitochondrial DNA sequences of various species of the genus Equus with special reference to the phylogenetic relationship between Przewalskii’s wild horse and domestic horse. The noncoding region between tRNAPro and the large conserved sequence block is the most variable region in the mammalian mitochondrial DNA D-loop region. This variable region (ca. 270 bp) of four species of Equus, including Mongolian and Japanese native domestic horses as well as Przewalskii's (or Mongolian) wild horse, were sequenced. These data were compared with our recently published Thoroughbred horse mitochondrial DNA sequences. The evolutionary rate of this region among the four species of Equus was estimated to be 2-4 x 10(-8) per site per year. Phylogenetic trees of Equus species demo...
Cloning and analysis of the cDNA encoding the horse and donkey luteinizing hormone beta-subunits. The coding regions of the horse (Equus caballus) and donkey (E. asinus) luteinizing hormone (LH) beta-subunit transcripts were cloned from pituitary gland RNA, in order to investigate their relationships to the corresponding equine chorionic gonadotropin (CG) beta-subunits and to further understand the unusual receptor-binding properties of equine LH and CG. The horse and donkey LH beta-subunit sequences were very similar (97% identity at the nucleotide (nt) level; 93% at the amino acid (aa) level), confirming their very close evolutionary linkage and also indicating that the C-terminal extens...
Incorporation of uracil into viral DNA correlates with reduced replication of EIAV in macrophages. The retrovirus equine infectious anemia virus (EIAV) encodes a dUTPase situated between reverse transcriptase and integrase. We have described the inability of EIAV with a 270-bp dUTPase deletion, delta DU EIAV, to replicate to wild-type (WT) levels in equine macrophages (D. S. Threadgill, W. K. Steagall, M. T. Flaherty, F. J. Fuller, S. T. Perry, K. E. Rushlow, S. F. J. LeGrice, and S. L. Payne, J. Virol. 67, 2592-2600, 1993). Here we describe the construction of a second dUTPase-deficient virus (DUD71E) containing a single amino acid substitution in dUTPase. delta DU and DUD71E replicate to ...
Complete primary sequence of equine cartilage link protein deduced from complementary DNA. Investigation of the structure of the equine articular cartilage link protein (LP) from individuals ranging in age from 1 to 15 years identified 3 distinct isoforms having molecular weights of 46,000, 43,000 and 41,000. The relative amounts of each of the 3 isoforms altered with age. The largest form did not change with age; however, amounts of the Mr 43,000 and 41,000 forms increased with increasing age. The results suggested that an accumulation, in the extracellular matrix of cartilage, of these 2 smaller products may have arisen from proteolytic cleavage. The complete amino acid sequence o...
The DNA sequence of equine herpesvirus 2. The complete DNA sequence of equine herpesvirus 2 (EHV-2) strain 86/67 was determined. The genome is 184,427 bp in size and has a base composition of 57.5% G + C. Unusually for a herpesvirus, about a third of the sequence distributed in several large blocks appears not to encode proteins. The 79 open reading frames that were identified as probably polypeptide-coding are predicted to encode 77 distinct proteins. Amino acid sequence comparisons confirmed that EHV-2 is a gamma-herpesvirus that is genetically collinear with herpesvirus saimiri (HVS; a gamma 2-herpesvirus) and Epstein-Barr virus (E...
Partial complementary deoxyribonucleic acid cloning of equine relaxin messenger ribonucleic acid, and its localization within the equine placenta. To determine the site of relaxin gene expression in equine placentae, a set of degenerate oligonucleotide primers was made according to the published amino acid sequence of the A- and B-chain of equine relaxin (eRXN). Total cellular RNA (tcRNA) from equine placentae at about 120 and 300 days of pregnancy was subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) with use of these primers. A single amplification product of approximately 430 bp was detected in each case by agarose gel electrophoresis. The PCR product was ligated into Bluescript plasmid and sequenced to confirm the...
Comparative ribotyping of Staphylococcus intermedius from dogs, pigeons, horses and mink. Strains of Staphylococcus (S.) intermedius from dogs, pigeons, horses and mink were typed by comparison of rRNA gene restriction fragment length polymorphisms (ribotyping) and the resulting ribotypes examined by cluster-analysis. Digestion of whole-cell DNA with HindIII resulted in 9 ribotypes with 3 to 4 bands. Separation of isolates from different host animal species was not possible. EcoRI yielded 11 different patterns with 4 to 9 fragments. The EcoRI-ribotypes of all canine strains grouped in one cluster encompassing four closely related ribotypes. Isolates were indistinguishable with resp...
At least four MHC class I genes are transcribed in the horse: phylogenetic analysis suggests an unusual evolutionary history for the MHC in this species. Nineteen horse MHC class I specificities have been serologically identified previously at a single locus (ELA-A), and two other specificities appear to be coded at other loci. Biochemical studies indicate that there are at least two expressed loci. In order to establish the number of transcribed horse MHC class I genes, we made a cDNA library from a heterozygous animal (ELA-A3/A7), and screened for positive clones using a bovine class I probe. More than 200 class I clones were isolated in this way, and so far seven unique full length sequences have been identified. All of the sequences are pre...
Extensive mtDNA diversity in horses revealed by PCR-SSCP analysis. The hypervariable D-loop region of mitochondrial DNA (mtDNA) was amplified with the polymerase chain reaction using total horse DNA samples. Analysis of single strand conformation polymorphism (SSCP) of denatured amplification products was carried out by native polyacrylamide (8%) gel electrophoresis followed by silver staining. As many as 15 distinct SSCP variants were revealed when screening a total of 78 maternally unrelated horses representing five different breeds. All breeds showed a high degree of polymorphism and the estimated probability (PImt) that two maternally unrelated individual...
Molecular cloning of cDNA for equine ovarian inhibin/activin beta A subunit. cDNAs encoding equine inhibin/activin beta A subunit were isolated from an equine follicle cDNA library and characterized. Using primers based on the rat inhibin/activin beta A subunit cDNA sequence, a RT-PCR was performed to generate the probe for screening. Four positive clones were isolated. Analysis of the nucleotide sequence of these clones revealed that two pairs of identical clones were present, Eq-beta A-1 (0.9 kb) and Eq-beta A-2 (1.5 kb). Eq-beta A-2 clone contained a complete open reading frame encoding 426 amino acids. The deduced amino acid sequence of equine inhibin/activin beta ...
Comparative RAPD-PCR analysis of lungworms (Dictyocaulidae) from fallow deer, cattle, sheep, and horses. Genomic DNA isolated from the four Dictyocaulus species D. viviparus, D. eckerti, D. filaria and D. arnfieldi was compared by random amplified polymorphic DNA polymerase chain reaction (RAPD)-PCR to get additional information whether lungworms from fallow deer belong to a separate species (D. eckerti) or have to be regarded as an isolate of D. viviparus in wild ruminants. The resulting banding patterns of the electrophoresed PCR products were compared to assess the degree of genetic differences between the different lungworms. For the two D. viviparus isolates a similarity coefficient of 93.4%...
Replication in vitro and in vivo of an equine infectious anemia virus mutant deficient in dUTPase activity. As an important enzyme in DNA synthesis, dUTPase is present in a wide variety of organisms and viruses and has been identified as a component of the equine infectious anemia virus (EIAV) pol gene. The role of EIAV dUTPase, designated DU, in virus replication in vitro and in vivo was investigated with a recently described infectious molecular clone of EIAV. A deletion mutant that was deficient in dUTPase activity was constructed, and its replication kinetics was examined in fetal equine kidney (FEK) cells and primary equine bone marrow macrophage (EBMM) cells. In FEK cells, which are permissive...
Regulatory function of the equine herpesvirus 1 ICP27 gene product. The UL3 protein of equine herpesvirus 1 (EHV-1) KyA strain is a homolog of the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1) and the ORF 4 protein of varicella-zoster virus. To characterize the regulatory function of the UL3 gene product, a UL3 gene expression vector (pSVUL3) and a vector expressing a truncated version of the UL3 gene (pSVUL3P) were generated. These effector plasmids, in combination with an EHV-1 immediate-early (IE) gene expression vector (pSVIE) and chimeric EHV-1 promoter-chloramphenicol acetyltransferase (CAT) reporter constructs, were used in trans...
Diversity of 16S rRNA genes of new Ehrlichia strains isolated from horses with clinical signs of Potomac horse fever. Ehrlichia risticii is the causative agent of Potomac horse fever. Variations among the major antigens of different local E. risticii strains have been detected previously. To further assess genetic variability in this species or species complex, the sequences of the 16S rRNA genes of several isolates obtained from sick horses diagnosed as having Potomac horse fever were determined. The sequences of six isolates obtained from Ohio and three isolates obtained from Kentucky were amplified by PCR. Three groups of sequences were identified. The sequences of five of the Ohio isolates were identical ...
Detection of Salmonella enteritidis in equine feces using the polymerase chain reaction and genus-specific oligonucleotide primers. Salmonella was identified in feces from horses, using the polymerase chain reaction (PCR) and genus-specific oligonucleotide primers. Feces from healthy horses were determined to be culture negative and PCR negative for Salmonella. Fecal samples were inoculated with known numbers of colony-forming units (CFU) of S. enteritidis. The fecal samples were enriched overnight in tetrathionate broth, and then DNA was extracted and amplified by PCR using genus-specific primers. Sensitivity of the assay extended to 10 degrees CFU Salmonella enteritidis/g feces; sensitivity of microbiologic culture with ...
Taxon-specific probes for the cellulolytic genus Fibrobacter reveal abundant and novel equine-associated populations. A total of six 16S rRNA targeted oligonucleotide probes were used to quantify Fibrobacter abundance and diversity in the gastrointestinal contents of a pony. Approximately 12% of the total 16S rRNA extracted from cecal contents hybridized with a Fibrobacter genus-specific probe and a Fibrobacter succinogenes species-specific probe. However, no significant hybridization was observed with a probe for the species. Fibrobacter intestinalis or with three probes for F. succinogenes subspecies. This suggested the presence of a previously undescribed population of F. succinogenes-like organisms. Novel...
