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Topic:Biotechnology
Biotechnology in horses encompasses the application of biological techniques and tools to enhance equine health, performance, and reproduction. This field includes genetic engineering, cloning, and the development of vaccines and therapeutics tailored to equine physiology. Techniques such as gene editing and stem cell therapy are explored for their potential to address genetic disorders, improve tissue regeneration, and enhance disease resistance in horses. Additionally, advancements in reproductive biotechnology, such as artificial insemination and embryo transfer, contribute to genetic diversity and breeding efficiency. This page compiles peer-reviewed research studies and scholarly articles that investigate the applications, methodologies, and implications of biotechnology in equine science.
Equine osteoclast-like cells generated in vitro demonstrate similar characteristics to directly isolated mature osteoclasts. We report on novel methods to isolate osteoclasts (OC s) and generate osteoclast-like cells (OCL s) from the bone and bone marrow of the equine femur. OC s were successfully isolated from bone scrapings taken from the endosteal surface of the femurs of three horses. OCL s were generated from bone marrow cells taken from the same animals. The validity of using the formation of OCL s as a method for studying OC differentiation and activity was confirmed by the similar characteristics of these two cells. In particular, they both were multinuclear, expressed the enzyme tartrate resistant acid phos...
Molecular cloning of equine transforming growth factor-beta1 reveals equine-specific amino acid substitutions in the mature peptide sequence. This study cloned and sequenced equine transforming growth factor (TGF)-beta1, yielding a unique nucleotide structure which predicted amino acid substitutions not seen in other mammalian species. The nucleotide sequence homology was 89% to bovine, 91% to man, 90% to ovine, and 86% to rat. Derived amino acid sequence comparison showed that the equine protein was unique, differing by two residues from man, cow, sheep, pig, and dog, and by three residues in the rat. Subsequent use of the cDNA clones to examine the expression of the TGF-beta1 gene in various tissues indicated predominant expressio...
Proton electron nuclear double resonance from nitrosyl horse heart myoglobin: the role of His-E7 and Val-E11. Electron nuclear double resonance (ENDOR) spectroscopy has been used to study protons in nitrosyl horse heart myoglobin (MbNO). (1)H ENDOR spectra were recorded for different settings of the magnetic field. Detailed analysis of the ENDOR powder spectra, using computer simulation, based on the "orientation-selection" principle, leads to the identification of the available protons in the heme pocket. We observe hyperfine interactions of the N(HisF8)-Fe(2+)-N(NO) complex with five protons in axial and with eight protons in the rhombic symmetry along different orientations, including those of the ...
Application of the restricted-access precolumn packing material alkyl-diol silica in a column-switching system for the determination of ketoprofen enantiomers in horse plasma. The group of LiChrospher ADS (alkyl-diol silica) sorbents that make part of a unique family of restricted-access materials, have been developed as special packings for precolumns used in the LC-integrated sample processing of biofluids. The advantage of these sorbents lies in the direct injection of untreated biological fluids, that is without sample clean-up, the elimination of the protein matrix with a quantitative recovery together with an on-column enrichment. The present method is based on previous work applying UV detection at 260 nm for ketoprofen determinations. Plasma samples introduc...
Generation of an equine oviductal epithelial cell line for the study of sperm-oviduct interactions. Equine oviductal epithelial cells (OEC) were transformed with simian virus 40 large T antigen (SV 40 T-ag) to create a cell line for the study of the interaction of equine spermatozoa with oviductal epithelium. One cell line was established based on the expression of the SV 40 T-ag and extended lifespan in culture. Immortalized equine OEC retained the characteristics of differentiated OEC such as the formation of monolayers with characteristic epithelial morphology and cell polarization as well as expression of cytokeratin and equine major histocompatibility complex I. Monolayers of immortaliz...
Synteny and regional marker order assignment of 26 type I and microsatellite markers to the horse X- and Y-chromosomes. The hypothesis that the conservation of sex-chromosome-linked genes among placental mammals could be extended to the horse genome was tested using the UCDavis horse-mouse somatic cell hybrid (SCH) panel. By exploiting the fluorescence in-situ hybridization (FISH) technique to localize an anchor locus, X-inactivation-specific transcript (XIST) on the horse X chromosome, together with the fragmentation and translocation of the X- and Y-chromosome fragments in a somatic cell hybrid panel, we regionally assigned 13 type I and 13 type II (microsatellite) markers to the horse X- and Y-chromosomes. T...
Molecular cloning and sequencing of the low-affinity IgE receptor (CD23) for horse and cattle. Expression of the low-affinity IgE receptor (CD23) on the surface of mononuclear cells is a critical event in the development of IgE-mediated immunologic responses. Using PCR and cDNA library screening, a 2.7kb cDNA encoding equine CD23 and a 627bp PCR fragment of cattle CD23 were sequenced and characterized. The equine CD23 sequence encodes a complete and continuous open reading frame of 327 amino acids, while the shorter cattle fragment encodes 209 amino acids corresponding to nucleotides 325-1098 of the equine CD23 transcript. In addition to high identities in their nucleotides and translat...
Isolation and characterization of an arterivirus defective interfering RNA genome. Equine arteritis virus (EAV), the type member of the family Arteriviridae, is a single-stranded RNA virus with a positive-stranded genome of approximately 13 kb. EAV uses a discontinuous transcription mechanism to produce a nested set of six subgenomic mRNAs from which its structural genes are expressed. We have generated the first documented arterivirus defective interfering (DI) RNAs by serial undiluted passaging of a wild-type EAV stock in BHK-21 cells. A cDNA copy of the smallest DI RNA (5.6 kb) was cloned. Upon transfection into EAV-infected BHK-21 cells, transcripts derived from this clo...
Comparison of sensitivity of sodium currents to tetrodotoxin in equine muscle specimens with that in murine and human muscle specimens. To determine sensitivity of equine skeletal muscle to tetrodotoxin and compare that with sensitivity of murine and human skeletal muscles. Methods: Semimembranosus, vastus lateralis, triceps brachii, and masseter muscle specimens from 22 euthanatized horses, vastus lateralis muscle biopsy specimens from 25 clinically normal humans, and diaphragmatic muscle specimens from 6 mice. Methods: Electrically elicited twitch responses were measured in muscle specimens incubated in medium alone and with tetrodotoxin (100 nM, 400 nM, 1.6 microM for equine specimens and 100 nM, 200 nM, 400 nM, 800 nM, 1.6...
Structure-related echoes in ultrasonographic images of equine superficial digital flexor tendons. To develop a method to discriminate between structure-related echoes and echoes resulting from interference, as observed in transverse ultrasonographic images of equine superficial digital flexor (SDF) tendons. Methods: 2 normal (injury-free) SDF tendons obtained from a 3-year-old Thoroughbred and a 9-year-old Dutch Warmblood horse. Methods: Tendons were mounted in a custom-made device that permitted exact transverse and perpendicular sequential scanning with precise steps of 0.5 mm along the long axis of the tendon. Photographs of transverse tendon sections at the exact scanning locations wer...
Assessment of muscle oxygenation in the horse by near infrared spectroscopy. This study examined the ability of near infrared spectroscopy (NIRS) to noninvasively determine changes to muscle oxygenation in the resting horse. Five horses had (NIRS) performed over extremity muscle while under general anaesthesia, first with 8 min limb ischaemia, then systemic hypoxaemia for 5 min. A second group of 6 awake horses had NIRS performed over extremity muscle while being administered hypoxic gas (F(I)O2 0.10) for 5 min, and after return to steady state, limb ischaemia was induced for an additional 5 min. In the anaesthetised horses' ischaemia induced marked and significant mus...
Modelling impact-initiated wave transmission through lung parenchyma in relation to the aetiology of exercise-induced pulmonary haemorrhage. Recently we proposed that exercise-induced pulmonary haemorrhage (EIPH) results from locomotory-impact-induced trauma by impact of the scapula on the chest wall during footfall and the consequent transmission of waves through the lung. A computational model has been developed to demonstrate that wave amplification and focusing occur in the dorsocaudal tip of the lung for waves originating on the anterior subscapular surface. The propagation of an acoustic wave was investigated in a simplified 2-dimensional representation of a vertical anterio-dorsal section of horse lung. It was demonstrated t...
A new method for quantitation of spin concentration by EPR spectroscopy: application to methemoglobin and metmyoglobin. A new method of EPR spectral analysis is developed to quantitate overlapping signals. The method requires double integration of a number of spectra containing the signals in different proportions and the subsequent solution of a system of linear equations. The result gives the double integral values of the individual lines, which can then be further used to find the concentrations of all the paramagnetic species present. There is no requirement to deconvolute the whole spectrum into its individual components. The method is employed to quantify different heme species in methemoglobin and metmyo...
Analyses of TCRB rearrangements substantiate a profound deficit in recombination signal sequence joining in SCID foals: implications for the role of DNA-dependent protein kinase in V(D)J recombination. We reported previously that the genetic SCID disease observed in Arabian foals is explained by a defect in V(D)J recombination that profoundly affects both coding and signal end joining. As in C.B-17 SCID mice, the molecular defect in SCID foals is in the catalytic subunit of the DNA-dependent protein kinase (DNA-PKCS); however, in SCID mice, signal end resolution remains relatively intact. Moreover, recent reports indicate that mice that completely lack DNA-PKCS also generate signal joints at levels that are indistinguishable from those observed in C.B-17 SCID mice, eliminating the possibilit...
Equine sarcoid tumour treated by autogenous tumour vaccine. Twenty-one horses with sarcoid tumours were treated by bio-immunotherapy using autogenous vaccines during 1991-1997. At operation the base of the tumour was left in the skin and autovaccines were made from extirpated tumour tissue by polymerization. The horses thus formed their own internal control group. One of 12 horses having a primary tumour, and four of 9 horses suffering recurrent tumours, prior to bio-immunotherapy, developed single recurrences. Four of these five horses suffering recurrence were treated anew, leading to remission. Disease-free intervals were longer for primary (P = 0.0...
Ultrastructure, protein phosphorylation and mRNA status of equine oocytes matured in vivo and in vitro. Equine oocytes were collected by follicle aspiration in vivo or by dissection of material obtained from an abattoir, and the ultrastructure, protein phosphorylation and mRNA status of the oocytes were evaluated. Electron microscopy studies indicated that the nucleus had a smooth membrane in oocytes with a compact cumulus, whereas the nuclear membrane was undulated in all other groups. Oocytes with compact cumuli had only a few microvilli, whereas those with expanded cumuli had more microvilli. There were only small numbers of cortical granules close to the oolemma in oocytes with compact cumul...
Indirect determination of stallion sperm capacitation based on esterase release from spermatozoa challenged with lysophosphatidylcholine. A spectrophotometric assay was developed to measure the amount of esterase released from stallion spermatozoa. This assay was used to determine the percentages of capacitated stallion spermatozoa, determined by the ability of spermatozoa to undergo an acrosome reaction and release esterase in response to a lysophosphatidylcholine challenge, for spermatozoa incubated under conditions to increase intracellular calcium and cAMP. Incubation with 100 nmol calcium ionophore A23187 l(-1) induced 66% of stallion spermatozoa to capacitate after 60 min of incubation at 37 degrees C. Subsequent experimen...
A comparative gene map of the horse (Equus caballus). A comparative gene map of the horse genome composed of 127 loci was assembled based on the new assignment of 68 equine type I loci and on data published previously. PCR primers based on consensus gene sequences conserved across mammalian species were used to amplify markers for assigning 68 equine type I loci to 27 horse synteny groups established previously with a horse-mouse somatic cell hybrid panel (SCHP, UC Davis). This increased the number of coding genes mapped to the horse genome by over 2-fold and allowed refinements of the comparative mapping data available for this species. In conju...
Quantitative real-time PCR for equine cytokine mRNA in nondecalcified bone tissue embedded in methyl methacrylate. Specific amplification and quantitation of nucleic acid sequences by the polymerase chain reaction (PCR) has been extensively used for the detection of viral infection and gene expression. Although successful amplification of DNA and RNA sequences extracted from paraffin embedded tissue have been described, there are presently no reports available regarding RNA analysis from bone and calcified tissues embedded in hydrophobic acrylic resin. Here we describe a general method for quantitation of specific mRNA sequences extracted from undecalcified bone sections, fixed in paraformaldehyde, and emb...
A DNA fragment of Leptospira interrogans encodes a protein which shares epitopes with equine cornea. Horses infected with Leptospira interrogans present several clinical disorders, one of them being recurrent uveitis. An antigenic relationship between this bacterium and equine cornea has been described in previous studies. With the aim to make progress on defining the molecular basis and pathogenesis of equine recurrent uveitis, here we describe the cloning of one DNA fragment from a Leptospira interrogans serovar pomona genomic lambda gt11 library. Although there are references of transcription of leptospiral genes in E. coli from their own leptospiral promoters, in this recombinant construc...
Cloning, expression, sequence analysis, and characterization of streptokinases secreted by porcine and equine isolates of Streptococcus equisimilis. Streptokinases secreted by nonhuman isolates of group C streptococci (Streptococcus equi, S. equisimilis, and S. zooepidemicus) have been shown to bind to different mammalian plasminogens but exhibit preferential plasminogen activity. The streptokinase genes from S. equisimilis strains which activated either equine or porcine plasminogen were cloned, sequenced, and expressed in Escherichia coli. The streptokinase secreted by the equine isolate had little similarity to any known streptokinases secreted by either human or porcine isolates. The streptokinase secreted by the porcine isolate had li...
Sequence and characterization of cDNA encoding the motilin precursor from chicken, dog, cow and horse. Evidence of mosaic evolution in prepromotilin. Motilin is involved in the regulation of the fasting motility pattern in man and in dog, but may have a different role in other species. Immunoreactive motilin has been demonstrated in several species, but the sequence is mostly unknown. The aim of this study was to isolate and sequence the cDNA encoding the motilin precursor from several mammalian species and from chicken. Total RNA was isolated from the duodenal mucosa of the chicken, dog, cow and horse. In each case single stranded cDNA was synthesized. Motilin cDNA fragments were amplified by PCR, ligated into a plasmid and cloned. Clones ...
The equine herpesvirus 2 E1 open reading frame encodes a functional chemokine receptor. Several herpesviruses contain open reading frames (ORFs) that encode potential homologs of eucaryotic genes. Equine herpesvirus 2 (EHV-2) is a gammaherpesvirus related to other lymphotropic herpesviruses such as herpesvirus saimiri and Epstein-Barr virus. The E1 ORF of EHV-2, a G protein-coupled receptor homolog, shows 31 to 47% amino acid identity with known CC chemokine receptors. To investigate whether E1 may encode a functional receptor, we cloned the E1 ORF and expressed it in stably transfected cell lines. We report here the identification of the CC chemokine eotaxin as a functional liga...
Equine synteny mapping of comparative anchor tagged sequences (CATS) from human Chromosome 5. Comparative anchor tagged sequences (CATS) from human Chromosome 5 (HSA5) were used as PCR primers to produce molecular markers for synteny mapping in the horse. Primer sets for 21 genes yielded eight horse-specific markers, which were mapped with the UC Davis horse-mouse somatic cell hybrid panel into two synteny groups: UCD14 and UCD21. These data, in conjunction with earlier human chromosome painting studies of the horse karyotype and synteny mapping of horse microsatellite markers physically mapped by FISH, confirm the assignment of UCD21 to ECA21 and suggest that UCD14 is located on ECA14...
Cloning and expression of a 48-kilodalton Babesia caballi merozoite rhoptry protein and potential use of the recombinant antigen in an enzyme-linked immunosorbent assay. A cDNA expression library prepared from Babesia caballi merozoite mRNA was screened with a monoclonal antibody BC11D against the rhoptry protein of B. caballi merozoite. A cDNA encoding a 48-kDa protein of B. caballi was cloned and designated BC48. The complete nucleotide sequence of the BC48 gene had 1,828 bp and was shown to contain no intron. Southern blotting analysis indicated that the BC48 gene contained more than two copies in the B. caballi genome. Computer analysis suggested that this sequence contained an open reading frame of 1,374 bp with a coding capacity of approximately 52 kDa. ...
The molecular genetics of red and green color vision in mammals. To elucidate the molecular mechanisms of red-green color vision in mammals, we have cloned and sequenced the red and green opsin cDNAs of cat (Felis catus), horse (Equus caballus), gray squirrel (Sciurus carolinensis), white-tailed deer (Odocoileus virginianus), and guinea pig (Cavia porcellus). These opsins were expressed in COS1 cells and reconstituted with 11-cis-retinal. The purified visual pigments of the cat, horse, squirrel, deer, and guinea pig have lambdamax values at 553, 545, 532, 531, and 516 nm, respectively, which are precise to within +/-1 nm. We also regenerated the "true" red ...
The equine herpes virus 4 thymidine kinase is a better suicide gene than the human herpes virus 1 thymidine kinase. The herpes simplex virus type 1 thymidine kinase suicide gene (HSV1tk) together with ganciclovir (GCV) have been successfully used for in vivo treatment of various experimental tumors, and many clinical trials using this system have been launched. With the aim to improve this therapeutic system, we compared the potential efficacy of different herpes virus derived thymidine kinases (HSV1, varicella-zoster virus, equine herpes virus type-4 and Epstein-Barr virus) as suicide genes in association with the nucleoside analogs acyclovir, ganciclovir and bromovinyldeoxyur- idine. Using various murine ...
