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.
The proximal ligand variant His93Tyr of horse heart myoglobin. The spectroscopic and structural properties of the His93Tyr variant of horse heart myoglobin have been studied to assess the effects of replacing the proximal His residue of this protein with a tyrosyl residue as occurs in catalases from various sources. The variant in the ferric form exhibits electronic spectra that are independent of pH between pH 7 and 10, and it exhibits changes in absorption maxima and intensity that are consistent with a five-coordinate heme iron center at the active site. The EPR spectrum of the variant is that of a high-spin, rhombic system similar to that reported for...
Diaphyseal structural properties of equine long bones. We evaluated the single-cycle structural properties for axial compression, torsion, and 4-point bending with a central load applied to the caudal or lateral surface of a diaphyseal segment from the normal adult equine humerus, radius, third metacarpal bone, femur, tibia, and third metatarsal bone. Stiffness values were determined from load-deformation curves for each bone and test mode. Compressive stiffness ranged from a low of 2,690 N/mm for the humerus to a high of 5,670 N/mm for the femur. Torsional stiffness ranged from 558 N.m/rad for the third metacarpal bone to 2,080 N.m/rad for the fe...
Validation of the shrinkage temperature of animal tissue for bioprosthetic heart valve application by differential scanning calorimetry. Shrinkage temperature is most often used to report the degree of cross-linking in glutaraldehyde-fixed animal tissue for use in bioprosthetic heart valve fabrication. Present practice utilizes the measurement of hydrothermal shrinkage observed when a sample is subjected to a temperature programme. This measurement at best gives a general indication of the efficiency of the treatment, i.e. the extent of cross-linking in the tissue. When differential scanning calorimetry has been used, the ambiguity arising from the scant reporting of the protocols used does not permit easy comparison of experim...
Cloning and expression of two genes from Babesia equi merozoites and evaluation of their diagnostic potential. High-titre equine immune sera were used to screen a lambda gt 11 expression library of Babesia equi cDNA fragments. Two cDNA clones which did not cross-hybridize to each other were studied. Both clones hybridized specifically to DNA from B. equi but not to DNA from B. caballi, B. divergens or B. ovis. Recombinant proteins were expressed as glutathione S-transferase (GST) fusion proteins with apparent molecular weights of 40 kDa and 75 kDa. Polyclonal antibodies directed against the 40 kDa and 75 kDa recombinant proteins detected native antigens of 55 kDa and 50 kDa respectively in crude lysate...
Electrostatics of hemoglobins from measurements of the electric dichroism and computer simulations. Hemoglobins from normal human cells, from sickle cells, and from horse were investigated by electrooptical methods in their oxy and deoxy forms. The reduced linear dichroism measured as a function of the electric field strength demonstrates the existence of permanent dipole moments in the range of 250-400 Debye units. The reduced limiting dichroism is relatively small (< or = 0.1); it is negative for hemoglobin from sickle cells and positive for the hemoglobins from normal human cells and from horse. The dichroism decay time constants are in the range from about 55 to 90 ns. Calculations of th...
Production of monoclonal antibodies in horses. Monoclonal antibodies (MAbs) have been successfully used to evaluate immune responses in horses, and to target important antigens of equine infectious agents to which protective immune responses may be directed (1–5). Most of these studies are performed with murine MAb produced by fusing spleen cells from immunized mice with an appropriate myeloma cell line, as described in Chapter 3. However, there are experimental questions for which murine MAb are not adequate. These include:
1.Definition of microbial epitopes recognized by the infected host;
2.Identification of immunodominant epito...
Nucleotide sequence of exons 5 to 9 of the p53 tumour-suppressor gene of the donkey (Equus asinus). The evolutionary conserved region of the equine homologue of the p53 gene from the donkey genome was PCR amplified and cloned. The 1380 bp fragment consisted of exons 5 to 9 and the intervening introns. The exonic and intronic DNA sequences showed a variable but high level of homology with previously published human sequences. The aminoacid sequences corresponding to the evolutionary conserved domains II, III, and V were identical to the human regions, whilst domain IV was 96% homologous.
Characterization of a density-corrected ultrasonic pneumotachometer for horses. A density-corrected ultrasonic pneumotachometer designed specifically for horses (UF202) was evaluated and characterized with the aid of a custom-built apparatus. UF202 provided voltage outputs for airflow through and gas density within the flowhead. Baseline stability for flow channel output (VUF202) was or = 0.9976). Under optimal conditions, VUF202 accuracy was determined to be +/- 1.00% FS and repeatability was +/- 0.78% FS. VUF202 resolution was 24 ml/s. The rise time for VUF202 was 18 ms, and the -3-dB point was 18 Hz; digital compensation provided a flat frequency response to 32 Hz. VU...
The identification of polymorphic microsatellite loci in the horse and their use in thoroughbred parentage testing. Six new horse microsatellite loci were identified by sequencing M13 clones containing horse genomic inserts which gave positive signals when probed with a CA/GT repeat probe. Oligonucleotide primer pairs were synthesized for these loci and for two previously described horse microsatellites, HTG4 and HTG6. Polymerase chain reaction assays were then carried out on a panel of 20 different unrelated Thoroughbred horse DNAs. DNAs from eight cases of double covering which could not be solved by conventional blood typing were also examined. Several of the loci amplified were found to be polymorphic a...
[Embryo transfer in horses–current status and future perspectives]. Although foals born after embryo transfer are eligible for registration in the majority of horse breeds, application of embryo transfer is still rare. This is mainly due to the lack of a possibility for superovulation. Uterine stage embryos can be recovered by a non-surgical flushing technique. Transfer can be accomplished by non-surgical as well as surgical methods. In contrast to the situation in cattle, most related technologies are scarcely available. Methods of cryopreservation as well as bisection of embryos are hampered by the fact that suitable embryos (morula) can be collected from th...
Recent developments in cryopreservation of stallion semen with special emphasis on thawing procedure using thermal hysteresis proteins. This research study explores the process of cryopreservation of stallion semen, focusing on improving the thawing procedures using thermal hysteresis proteins (THPs) from Antarctic and Arctic fish in order to […]
Equilibrium unfolding studies of horse muscle acylphosphatase. The stability and equilibrium unfolding behaviour of horse muscle acylphosphatase have been studied by denaturing the protein under various conditions of temperature, pH, and urea concentration. Far-ultraviolet circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy indicate that this small monomeric protein unfolds reversibly and cooperatively. Thermodynamic parameters, the Gibbs free energy delta G and enthalpy delta H of unfolding, have been estimated for denaturation of the protein from NMR and CD data as 19 kJ mol-1 and 350 kJ mol-1, respectively. CD and 1H-NMR results s...
Genetic Bit Analysis: a solid phase method for typing single nucleotide polymorphisms. A new method for typing single nucleotide polymorphisms in DNA is described. In this method, specific fragments of genomic DNA containing the polymorphic site(s) are first amplified by the polymerase chain reaction (PCR) using one regular and one phosphorothioate-modified primer. The double-stranded PCR product is rendered single-stranded by treatment with the enzyme T7 gene 6 exonuclease, and captured onto individual wells of a 96 well polystyrene plate by hybridization to an immobilized oligonucleotide primer. This primer is designed to hybridize to the single-stranded target DNA immediately...
In vitro fertilization rate of horse oocytes with partially removed zonae. Frozen-thawed ejaculated stallion spermatozoa were preincubated for 3 h in BO medium containing 5 mM caffeine and then treated with 0.1 micro M calcium ionophore A23187 for 60 sec. Aliquots of the sperm suspension (final concentration 1-2 x 10(7)/ml) were added to the oocytes which had been matured in vitro for 32 h. In Experiment 1, there were 3 groups of oocytes; cumulus intact, denuded zona-intact, and zona-free. Cumulus cells were removed with 0.5% hyaluronidase and the zona pellucida with 0.1% protease. The oocytes were fixed 20 h after insemination with acetic acid:ethanol (1:3) and stai...
Molecular cloning and expression of two horse pancreatic cDNA encoding colipase A and B. Pancreatic colipase plays an essential role in the intestinal fat digestion by anchoring lipase on lipid/water interfaces in the presence of bile salts. In contrast to other species, two molecular forms of colipase, A and B, have been found in horse. The two corresponding cDNAs were isolated from a horse pancreatic library and their nucleotide sequences were determined. Moreover, for the first time, active colipase has been obtained after transfection of COS cells by either colipase A or B cDNA.
Polymorphic sequence in the D-loop region of equine mitochondrial DNA. The D-loop regions in equine mitochondrial DNA were cloned from three thoroughbred horses by polymerase chain reaction (PCR). The total number of bases in the D-loop region were 1114 bp, 1115 bp and 1146 bp. The equine D-loop region is A/T rich like many other mammalian D-loops. The large central conserved sequence block and small conserved sequence blocks 1, 2 and 3, that are common to other mammals, were observed. Between conserved sequence blocks 1 and 2 there were tandem repeats of an 8 bp equine-specific sequence TGTGCACC, and the number of tandem repeats differed among individual horses....
Molecular dynamics simulation of equine infectious anemia virus Tat protein in water and in 40% trifluoroethanol. Two molecular dynamics (MD) simulations were performed in order to increase the understanding of the dependence of protein conformation on solvent environment. The protein used for these simulations is the transcriptional activator of the equine infectious anemia virus (EIAV-Tat). The structure of this protein has been determined by nuclear magnetic resonance (NMR) in aqueous solution (Willbold et al., Science 264, 1584 (1994)) and in 40% (v/v) trifluoroethanol (TFE) (Sticht et al., Eur. J. Biochem., submitted) showing considerable differences in the stability of the secondary structure elemen...
Models of the three-dimensional structures of echidna, horse, and pigeon lysozymes: calcium-binding lysozymes and their relationship with alpha-lactalbumins. Similarities in amino acid sequences, three-dimensional structures, and the exon-intron patterns of their genes have indicated that c-type lysozymes and alpha-lactalbumins are homologous proteins, i.e., descended by divergent evolution from a common ancestor. Like the alpha-lactalbumins, echidna milk, horse milk, and pigeon eggwhite lysozymes all bind Ca(II). Models of their three-dimensional structures, based on their amino acid sequences and the known crystal structures of domestic hen eggwhite and human lysozymes and baboon and human alpha-lactalbumins, have been built. The several structur...
Differential scanning calorimetric studies of superficial digital flexor tendon degeneration in the horse. Differential scanning calorimetry (DSC) of equine superficial digital flexor tendons revealed the presence of a small exothermic peak at 23 degrees C of unknown origin, and a large endothermic peak at 70 degrees C due to denaturation of cross-linked collagen fibres. In the central degenerated core of damaged tendons the denaturation temperature remained at 70 degrees C but the enthalpy decreased in relation to the extent of degeneration of the tendon. We suggest that this reduction in enthalpy is due to depolymerisation and denaturation of the collagen fibres. This contention is supported by t...
FTIR analysis of the interaction of azide with horse heart myoglobin variants. The interaction of azide with variants of horse heart myoglobin (Mb) has been characterized by Fourier transform infrared (FTIR), electron paramagnetic resonance (EPR), and UV-VIS absorption spectroscopy and by molecular modeling calculations. Distal histidine variants (His64Thr, His64Ile, His64Lys) and charged surface variants (Val67Arg, Lys45Glu, Lys45Glu/Lys63Glu) were included in this study. All variants, with the exception of Val67Arg, have a lower azide affinity than the wild-type protein. Analysis of the temperature dependence of the FTIR spectra (277-313 K) revealed that the wild-type ...
Sequence of a cDNA encoding horse growth hormone. A cDNA encoding horse growth hormone (ecGH) was isolated and sequenced. The coding sequence resembles a typical mammalian GH pre-protein and contains a 3' untranslated region of 101 nucleotides carrying two contiguous polyadenylation signals.
Identification of the horse epidermal growth factor (EGF) coding sequence and its use in monitoring EGF gene expression in the endometrium of the pregnant mare. The PCR technique and highly degenerate oligonucleotide primers were used to amplify a 282 bp fragment of the horse (Equus caballus) epidermal growth factor (EGF) cDNA. The clone corresponded to 94 amino acids of the EGF precursor molecule. The deduced amino acid sequence of the 53 residue EGF mitogenic peptide within the precursor sequence showed 60-70% identity with five other published EGF sequences. The PCR cDNA fragment hybridized to a 4.9 kb transcript in horse kidney and endometrial RNA which was of a similar size to the mature EGF transcript found in other mammalian species. The horse ...
Cloning and sequencing of the equine testicular follitropin receptor. To investigate the possibility that specific structural determinants within the equine follitropin receptor (eFSHR) are critical to the enhanced specificity of this receptor compared to other FSHRs, we used the RACE-PCR technique to clone the eFSHR from equine testis. Sequence analysis revealed that the eFSHR is highly homologous to other mammal FSHRs, but it presents 10 unique amino acid residue replacements in the extracellular domain. Furthermore, a potential N-glycosylation site was detected at a position not encountered in other receptors. Northern blot analysis identified three transcrip...
Purification of a plasminogen activator from Streptococcus uberis. A protein capable of activating bovine, equine and ovine plasminogen, but not that from human or porcine plasma, was purified from culture filtrates of Streptococcus uberis (strain 0140J). Purification was achieved by ammonium sulphate precipitation followed by molecular exclusion chromatography. The elution position of the native molecule was equivalent to a molecular mass of approximately 57 kDa. However, the molecular mass, as determined by SDS-PAGE, was 29 kDa, suggesting the existence of a dimeric structure. Purified immunoglobulin from three out of five monoclonal antibodies raised to th...
Studies on glycoprotein-derived carbohydrates. This research focuses on the study of glycoproteins, specifically investigating their carbohydrate chains and their various functions in living organisms. The article highlights the challenges in isolating specific carbohydrate chains […]
The transition from inhomogeneous to homogeneous kinetics in CO binding to myoglobin. Heme proteins react inhomogeneously with ligands at cryogenic temperatures and homogeneously at room temperature. We have identified and characterized a transition from inhomogeneous to homogeneous behavior at intermediate temperatures in the time dependence of CO binding to horse myoglobin. The turnover is attributed to a functionally important tertiary protein relaxation process during which the barrier increases dynamically. This is verified by a combination of theory and multipulse measurements. A likely biological significance of this effect is in the autocatalysis of the ligand release p...
Rapid refolding of native epitopes on the surface of cytochrome c. Refolding of surface epitopes on horse cytochrome c has been measured by monoclonal antibody binding. Two antibodies were used to probe re-formation of native-like surface structure: one antibody (2B5) binds to native cytochrome c near a type II turn (residue 44) while the other (5F8) binds to a different epitope on the opposite face of the protein near the amino terminus of an alpha-helical segment (residue 60). The results show that within the first approximately 100 ms of refolding all of the unfolded protein collapses to native-like folding intermediates that contain both antibody binding ...