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.
Molecular cloning of cDNA for equine follistatin and its gene expression in the reproductive tissues of the mare. A cDNA clone encoding equine follistatin was isolated from an equine ovarian cDNA library. Out of 1.2 x 10(5) independent clones screened, one positive clone was isolated and its cDNA sequence determined. The isolated clone, named EQ-FS-1, contained a complete open reading frame encoding 344 amino acid residues. The similarity of its deduced amino acid sequence to these of other mammalian species was greater than 95%. Although its expression level varied among the tissues examined, follistatin mRNA was detected in the equine uteroplacental tissues, follicles and corpora lutea by Northern blot ...
Antigenic profile of African horse sickness virus serotype 4 VP5 and identification of a neutralizing epitope shared with bluetongue virus and epizootic hemorrhagic disease virus. African horse sickness virus (AHSV) causes a fatal disease in horses. The virus capsid is composed of a double protein layer, the outermost of which is formed by two proteins: VP2 and VP5. VP2 is known to determine the serotype of the virus and to contain the neutralizing epitopes. The biological function of VP5, the other component of the capsid, is unknown. In this report, AHSV VP5, expressed in insect cells alone or together with VP2, was able to induce AHSV-specific neutralizing antibodies. Moreover, two VP5-specific monoclonal antibodies (MAbs) that were able to neutralize the virus in a ...
Structure of mare apolactoferrin: the N and C lobes are in the closed form. The structure of mare apolactoferrin (MALT) has been determined at 3. 8 A resolution by the molecular-replacement method, using the structure of mare diferric lactoferrin (MDLT) as the search model. The MDLT structure contains two iron-binding sites: one in the N-terminal lobe, lying between domains N1 and N2, and one in the C-terminal lobe between domains C1 and C2. Both lobes have a closed structure. MALT was crystallized using the microdialysis method with 10%(v/v) ethanol in 0.01 M Tris-HCl. The structure has been refined to a final R factor of 0.20 for all data to 3.8 A resolution. Compar...
Construction of chromosome-specific paints for meta- and submetacentric autosomes and the sex chromosomes in the horse and their use to detect homologous chromosomal segments in the donkey. A pilot study comparing horse and donkey karyotypes on a molecular basis was initiated using the chromosomal microdissection approach. All equine meta- and submetacentric chromosomes, viz. ECA1 to ECA13 and the X and Y chromosomes, were microdissected. The DNA was PCR amplified, non-radioactively labelled and used as probes on equine metaphase chromosomes to confirm their origin. Once tested, the paints were used as probes on donkey metaphase chromosomes to detect homologous chromosomal segments between the two species. The results not only detected conservation of whole chromosome and/or arm ...
Identification of a new aspartic proteinase expressed by the outer chorionic cell layer of the equine placenta. The pregnancy-associated glycoproteins (PAGs) are placental antigens that were initially characterized as pregnancy markers in the maternal circulation of domestic ruminant species. They are members of the aspartic proteinase gene family, having greatest sequence identity with pepsinogens. However, some are not capable of functioning as enzymes. The PAGs are associated with a large gene family within the Artiodactyla order (cattle, camels, pigs). So far, no members of this family have been characterized in species outside this order. This report describes the cloning and initial characterizati...
Molecular dynamics simulation of alpha-lactalbumin and calcium binding c-type lysozyme. Alpha-lactalbumins (LAs) and c-type lysozymes (LYZs) are two classes of proteins which have a 35-40% sequence homology and share a common three dimensional fold but perform different functions. Lysozymes bind and cleave the glycosidic bond linkage in sugars, where as, alpha-lactalbumin does not bind sugar but participates in the synthesis of lactose. Alpha-lactalbumin is a metallo-protein and binds calcium, where as, only a few of the LYZs bind calcium. These proteins consist of two domains, an alpha-helical and a beta-strand domain, separated by a cleft. Calcium is bound at a loop situated at...
Extraction of equine chorionic gonadotrophin from pregnant mare plasma by direct adsorption on chromatographic media. Equine chorionic gonadotrophin (eCG) is a hormone of practical value in veterinary medicine and animal production. Here we report a novel preparation procedure based on its direct adsorption onto anionic-exchange resins in a batch-wise mode. The active plasma is previously conditioned to reduce pH and ionic strength to required levels. After the adsorption stage, a 90% recovery of the initial eCG is achieved, with a concentration factor of about 50 and an enrichment factor around 500, with high preservation of biological activity. Further purification is carried out by cation-exchange column c...
Presence and comparison of angiotensin converting enzyme in commercial cell culture sera. This study was conducted to determine the presence of the angiotensin converting enzyme in commercial sera used in cell culture medium. The aim of the research was to bring the presence of proteinases (angiotensin converting enzyme) to cell culture users' knowledge and to give some data for solving problems about the development of peptides as useful drugs. The enzymes, purified from foetal bovine, adult bovine, foetal equine, adult equine, and human sera, showed molecular weights of about 170 kDa. Captopril and lisinopril inhibited enzyme activities at nanomolar concentrations. The enzymes we...
Comparative mapping of 18 equine type I genes assigned by somatic cell hybrid analysis. Polymerase chain reaction primers designed from horse cDNA sequences and from consensus sequences highly conserved in mammalian species were used to amplify markers for synteny mapping 18 equine type I genes. These markers were used to screen a horse-mouse somatic cell hybrid panel (UCDavis SCH). Fourteen primer sets amplified horse-specific fragments, while restriction enzyme digests of PCR products were used to distinguish the fragments amplified from horse and mouse with four primer sets. Synteny assignments were made based on correlation values between each marker tested and other markers ...
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 gr...
A technique for magnetic resonance imaging of equine cadaver specimens. We tested an adaptation of a technique for performing magnetic resonance (MR) imaging of human cadaver limbs in the horse. The forelimbs from a normal horse were collected, frozen, and sealed with a paraffin-polymer combination prior to imaging with either a high- or midfield magnetic resonance scanner. Each forelimb was defrosted, scanned, and refrozen on two separate occasions. A five-point scale was used to evaluate the quality of each set of sagittal and transverse, T1-weighted images of each digit. There was no difference in image quality between first and second scans of either specimen ...
Quantitative measurement of equine cytokine mRNA expression by polymerase chain reaction using target-specific standard curves. Quantification of cytokine mRNA using reverse transcription coupled with the polymerase chain reaction (RT-PCR) has become a corner stone of the study of cytokine regulation. Quantitative competitive RT-PCR (QCRT-PCR) is commonly accepted as a reliable method for quantifying differences in mRNA levels but is both labor- and reagent-intensive. A noncompetitive polymerase chain reaction method that utilizes cytokine-specific, plasmid-derived, standard curves was developed for the quantification of equine cytokine mRNA. The assay can be performed on minute samples of cellular material, utilizes s...
Cytokeratins of the equine hoof wall, chestnut and skin: bio- and immunohisto-chemistry. The equine skin and its appendages (chestnut, hoof capsule, ergot, sebaceous glands, sweat glands and hair) consist mainly of keratinocytes. The intermediate filament cytoskeleton of these cells in involved in specialised functions, such as mechanical co-ordination of the cytoskeleton of the cell or tissue. In this study, 7 monoclonal antibodies, one polyclonal antibody and immunoblot analysis were used to characterise cytokeratins (separated by 1- and 2-dimensional gel electrophoresis) from the hoof wall and chestnut. The tissue distribution of these cytokeratins was studied by immunohistoche...
Human chorionic gonadotropin induces an inverse regulation of steroidogenic acute regulatory protein messenger ribonucleic acid in theca interna and granulosa cells of equine preovulatory follicles. The time- and gonadotropin-dependent regulation of steroidogenic acute regulatory protein (StAR) has not been characterized in vivo in preovulatory follicles of large monoovulatory species or sexually mature animals. The objectives of this study were to clone equine StAR and describe the regulation of its messenger RNA (mRNA) in equine follicles after the administration of an ovulatory dose of hCG. The screening of an equine follicle complementary DNA (cDNA) library with a mouse StAR cDNA probe revealed two forms of equine StAR that differ only in the length of their 3'-untranslated region (3'...
Micromechanics of the equine hoof wall: optimizing crack control and material stiffness through modulation of the properties of keratin. Small-scale components of the equine hoof wall were tested to determine their mechanical roles in the morphological hierarchy. Macroscale tensile tests conducted on samples of the inner wall tubules and intertubular material showed a sixfold difference in mean initial stiffnesses (0.47 and 0.08 GPa, respectively), indicating that the inner wall tubules stiffen the wall along its longitudinal axis. The similarity in material properties of tubule and intertubular samples from the mid-wall suggests that tubules in this region offer only minor reinforcement along the longitudinal axis. Microscale ...
Molecular characteristics of equine stromelysin and the tissue inhibitor of metalloproteinase 1. To clone the entire coding sequence of equine matrix metalloproteinase-3 (MMP-3, stromelysin) and tissue inhibitor of metalloproteinase-1 (TIMP-1) and compare their nucleotide and amino acid sequences with those of MMP-3 and TIMP-1 from other species. Methods: Articular cartilage harvested from the joints of 4 foals, 2 yearlings, and 3 adult horses. Methods: A cDNA library was constructed from mRNA extracted from equine chondrocytes. The library was screened and clones selected that contained the cDNA for MMP-3 and TIMP-1. The cDNA was sequenced and the nucleotide and deduced amino acid sequen...
The 2.03 signal as an indicator of dinitrosyl-iron complexes with thiol-containing ligands. The parameters of EPR signal from dinitrosyl-iron complexes (DNIC) with bovine serum albumin (BSA), horse hemoglobin (Hb), and apometallothionein (apo-Mt) of horse kidney incorporating one (BSA, Hb) or two thiol-containing ligands (apo-Mt) were compared. The EPR signal from DNIC-BSA was characterized by the rhombic symmetry of g tensor at room temperature of signal recording (ambient temperature) or at 77K in the solution frozen in the presence of glycerol. In freezing of the solution in the absence of glycerin, under the exposure of DNIC-BSA to negatively charged sodium dodecyl sulfate (SDS) ...
Relative binding of therapeutic drugs by sera of seven mammalian species. The relative binding of acetaminophen, lidocaine, phenobarbital, procainamide, quinidine, and theophylline to sera of seven mammalian species was studied. Pooled commercial sera from cow, goat, horse, human, pig, rabbit, and sheep were supplemented with 5 and 10 mM concentrations of each drug. For each serum, each drug, and each drug concentration, equilibrium dialysis was performed in duplicate against phosphate buffer (pH 7.4, 0.1 M, 4 degrees C). Percent drug bound to serum was calculated. Phenobarbital demonstrated more than 20% binding to goat, horse, human, and sheep serum at both 5 and ...
In vitro generation of equine osteoclasts from bone marrow cells using a novel culture system. We report on preliminary results of a novel in vitro culture system designed to generate equine osteoclasts in large numbers. Osteoclast generation, as determined by the expression of tartrate resistant acid phosphatase (TRAP) and ability to resorb bone, was enhanced in equine bone marrow cultures supplemented with fibroblastic cell (L929) conditioned medium (L929-CM). Bone marrow was collected from a total of 12 horses and ponies and TRAP-positive cells with bone resorbing ability were generated in significant numbers in the last seven. TRAP-positive mononuclear cells appeared after three day...
Cloning and characterization of the equine F18 gene, which has a novel exon. A genomic clone isolated from an equine genomic library probed with an oligonucleotide (CAG)10 showed high sequence similarity to the human F18 gene and was tentatively named equine F18 gene. Because the human F18 gene is expressed in many tissues, we examined whether this equine clone was also expressed in equine tissues. The cDNA encoding equine F18 was obtained by the reverse transcriptase-polymerase chain reaction (RT-PCR) from equine thymus. The nucleotide sequence of the equine F18 cDNA (1940 bp) was determined and contained both the ATG initiation codon and a poly(A) sequence. The cDNA ...
Pressure and temperature dependence of enantioselective excited-state quenching of chiral Tb(III) and Eu(III) tris(pyridine-2,6-dicarboxylate) chelates by various C-type ferricytochromes. For mitochondrial ferricytochrome c from horse, cow and tuna and for bacterial cytochrome c-550 from Paracoccus versutus, the pressure and temperature dependence of their quenching of racemic Tb(DPA)3(3-) and Eu(DPA)3(3-) (DPA = pyridine-2,6-dicarboxylate) luminescence in aqueous solution is investigated. Of these energy transfer reactions the activation volumes (delta V#) and energies (Ea) are determined for the ranges P = 0-3 kbar and T = 15-40 degrees C. For the lambda enantiomers of Tb(DPA)3(3-) and Eu(DPA)3(3-), delta V# and Ea are almost the same for all proteins: 0.4 < or = delta V# < o...
Equilibrium and kinetics of the folding of equine lysozyme studied by circular dichroism spectroscopy. The equilibrium unfolding and the kinetics of unfolding and refolding of equine lysozyme, a Ca2+-binding protein, were studied by means of circular dichroism spectra in the far and near-ultraviolet regions. The transition curves of the guanidine hydrochloride-induced unfolding measured at 230 nm and 292.5 nm, and for the apo and holo forms of the protein have shown that the unfolding is well represented by a three-state mechanism in which the molten globule state is populated as a stable intermediate. The molten globule state of this protein is more stable and more native-like than that of alp...
Monitoring the conformational flexibility of cytochrome c at low ionic strength by 1H-NMR spectroscopy. Horse heart cytochrome c at pH 7 and low ionic strength is present as two conformers, as evidenced by 1H-NMR spectroscopy. The two structures have been calculated using NOE and pseudocontact shift constraints. They have the same folding patterns and are essentially equal, within the rmsd of the families. The two average structures have rmsd values of 0.049 nm and 0.093 nm for the backbone and the heavy atoms, respectively. Such a difference has been analyzed through a detailed analysis of the NOEs. It appears that the species at low ionic strength differs from the species present at high ionic...
Controlled-release products for the control of the estrus cycle in cattle, sheep, goats, deer, pigs, and horses. This paper describes the estrus cycles of a number of livestock breeds and reviews the controlled-release drug delivery systems that are currently available for the purpose of controlled breeding. The bovine estrus cycle is reviewed in detail, and the estrus cycles of other species are described in a manner that highlights similarities and differences between species. Pertinent formulation and pharmacokinetic information about current drug delivery systems is presented and discussed, and recent advances in this area are also described.
Isolation and chondrocytic differentiation of equine bone marrow-derived mesenchymal stem cells. To isolate mesenchymal stem cells from adult horses and determine specific monolayer culture conditions required to enhance biochemically and phenotypically defined chondrocytic differentiation. Methods: 2 adult horse bone marrow donors without skeletal or hematologic abnormalities. Methods: Bone marrow was aspirated from the sternebra, and mesenchymal stem cells were isolated by centrifugation and cultured in monolayers. Subcultures were established in 24-well plates on day 13. Culture medium was harvested every 2 days, and culture of 12 of the 24 wells was terminated on day 6 and of the rema...