_INSTITUTE OF ANIMAL BREEDING & GENETICS _Image information

 

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_IVP-ET twins In vitro production (IVP) and embryo transfer (ET) are basic techniques in biomedicine and biotechnology. We have developed a method, based on endoscopy, for the collection and transfer of embryos. The picture shows cattle twins, produced by this techniques, with the recipient mother.

_CsCl gradient Centrifugation of DNA in CsCl solution results in the formation of a density gradient defined by the GC-content of the DNA. A central band is formed by the main fraction of DNA in the centrifugation tube. In the presence of ethidium bromide, the DNA band shows fluorescence under UV illumination.

_Blastocyst Blastocysts represent an embryonic stage in mammalian development, consisting of a hollow ball of surface cells and an inner cell mass. Blastocysts are used in different experiments in biomedicine and biotechnology.

_Obese mice Leptin receptor (Lepr, Ob-R) knock-out mice show severe early-onset obesity and extreme insulin resistance. An affected six month-old female is shown together with a control littermate. In addition to this previously described obesity phenotype, we observed homozygous leptin receptor mutant mice developing cachexia after having shown initial obesity.

_DNA DNA (Deoxyribonucleic acid) encodes the proteins and their specific spatio-temporal expression in the cell. The first step in the molecular genetic characterization of biological material is the isolation of the DNA. A mammalian DNA precipitate in ethanol is shown.

_GFP transfected cells Transfection is the commonly used procedure for analyzing mammalian gene expression in vivo. The parameters for transfection differ nearly for every cell line, therefore, it is necessary to optimize these parameters. The green fluorescent protein (GFP) is used for these experiments, because it serves as an easily detectable fluorescent tag requiring no substrates or associated factors.

_California rabbit Temperature-sensitive melanisation was observed in California rabbits resulting in the exclusive pigmentation of the extremities. Cooling of unpigmented parts of the coat induces pigmentation. This phenomenon is caused by temperature-sensitive tyrosinase alleles and has been also described in cats, humans and 'himalayan' mice.

_MHS diagnosis The malign hyperthermic syndrome (MHS) in pig is caused by a defined point mutation in the RYR1 gene. Molecular genetic testing by PCR and subsequent restriction enzyme digestion reveals the three different MHS genotypes after gel electrophoresis: PP - one band, NN - two bands, NP - three bands.

_GFP-mice We have produced transgenic mice expressing the green fluorescent protein (GFP, originally from the jellyfish Aequorea victoria) under the control of various promoter elements. Transgenic mice expressing GFP driven by tissue specific promoters contribute to the investigation of embryogenesis.

_Microinjection Transgenes are microinjected into the pronuclei of fertilized oocytes and will be randomly integrated within the host genome. Developing embryos are transfered into the oviduct of recipient animals. Some of the resulting offspring show germ line integration of the transgene and serve for the generation of transgenic lines.

_Cattle clones Cloned calves (Simmental breed) produced by nuclear transfer of an embryonic donor cell into a recipient cytoblast. The embryonic donor cell derived from a 92-cell donor morula.

_MS chromatogram After electrophoretical separation of fluorescence labelled microsatellite (MS) PCR amplificates, the fragment length is automatically analyzed by suitable software. The chromatogram shows one microsatellite locus of a heterozygous animal, whose alleles differ in their length by 4 base pairs.

_KBR Polymorphic surface epitopes on red blood cells are traditionally used as identity markers. In cattle, 60 different blood markers are tested for presence or absence by monovalent antisera in the komplement binding reaction (KBR). The image shows positive and negative KBRs, whereby  the positive reaction is determined by haemolysis.

_Mongrel Mongrels are said to be healthier and more intelligent than purebred dogs as a consequence of their higher heterozygosity. In our studies, we found no differences in the prevalence of diseases in purebred dogs with high inbreeding level and in mongrels.

_CEA The collie eye anomaly (CEA) is an autosomal recessive defect. The picture shows a severe chorioideo-retinal  hypoplasia and a coloboma of the optic disc, at 7 o'clock position.

_Hip dysplasia Hip dysplasia is a congenital disease that affects large dog breeds. It causes weakness and dysfunction of the back quarters and leads to painful arthritis. Outbreak of the disease is influenced by environmental factors. Today hip dysplasia is diagnosed by radiography.

_Metaphase spreads Metaphase chromosomes are spread on slides and labeled with DNA binding dyes. The resulting species specific banding pattern is used to identify and characterize individual chromosomes.

_Persian cat Persian cats are prone to eye diseases as conjunctivitis as a consequence of the short nose and the tight nasolacrimal ducts.  In our studies a higher number of red blood cells was observed to compensate the decrease of oxygen uptake that is caused by the narrowed upper respiratory tract.

_FISH analysis Fluorescence in situ hybridization (FISH) is used to localize target nucleotide sequences within tissues, cells, intracellular structures and on individual chromosomes. Detection is carried out by complementary probes conjugated to a fluorescent compound.

_Real time PCR The method of real-time PCR has been developed to measure input target templates quantitatively. PCR quantitation has been used in a wide variety of applications, including the evaluation of gene copy number and mRNA expression, the diagnosis and management of infectious diseases, to investigate the efficiency of gene delivery systems, to verify biological product purity and to qualitatively detect genes and alleles in high-throughput assays.

_DNA-microsatellites DNA microsatellites are highly polymorphic, abundantly present and selectively neutral markers in the genome. Therefore microsatellites are ideal identity markers. After carrying out fluorescence labeled multiplex PCR, the amplificates are electrophoretically separated and automatically detected. The gel image shows the results for 11 different bovine microsatellite loci applied per lane.

_Trace material Forensic trace material differs from case to case. The picture shows a blood stain in the snow. This trace material was sent to us by a huntsman who wanted to know, what species of game has been poached in his hunting ground. The evidence, it was a roa deer, was supplied by molecular genetic species identification.

_Southern analysis The principle of the Southern hybridization is that a single-stranded DNA molecule of  a defined sequence ("probe”) base-pairs to a second DNA molecule containing the complementary sequence ("target”). Therefore, the target DNA is electrophoretically separated and transferred to a membrane, where hybridisation with tha labeled probe takes place. This analysis is very sensitive and permits detection of single-copy genes in complex genomes.

_Blue white selection The blue/white color screening is used to identify recombinant clones (transformants) in cloning experiments. Cloning vectors produce ß-galactosidase by complementation when transformed into bacteria. This causes the cells to exhibit a blue colour in the presence of the substrate X-Gal. Foreign DNA inserted in the cloning vector disrupts the lacZ gene, which prevents complementation and results in the appearance of white colonies.

_Sequencing analysis DNA sequence determination is an encymatic method where DNA fragments of different length are produced. Base specific fluorescence labeling of the fragments enables the detection and discrimination of the fragments by laser light during polyacrylamide gel electrophoresis.

_Laboratory mice The mouse is the most widely used animal in biomedical research. The laboratory mouse, Mus domesticus domesticus originated from wild house mice bred in the nineteenth century. Inbred lines are usually used in the experiments because of their genetic uniformity.

_Embryonic stem cells Embryonic stem cells (ES cells) are pluripotent cells derived from the inner cell mass of blastocysts. Under defined culture conditions, these cells maintain their embryonic developmental potential even after many passages of in vitro culture. Following genetic manipulation and subsequent introduction into an embryo, they are able to successfully colonize every cell lineage including germ line.

_Blastocyst microinjection Embryonic stem cells are used for knockout experiments and, therefore, are transfected in vitro with a gene construct. After homologous recombination in the ES cell genome the ES cells are injected into blastocysts. Subsequently the injected blastocysts are transfered to recipient animals. Some of the resulting offsprings will have the transgene integrated into the germ line and can serve for the production of transgenic lines.

_Blood samples Blood-typing is done on intact red blood cells by komplement binding reaction (KBR) and by blood protein electrophoresis. DNA genotyping is done on blood samples as well as on other tissues (sperm, saliva, skin, hair bulbs).

_PAGE of blood proteins Blood proteins are genetically polymorphic. Individual variants are separated in native polyacrylamide gel electrophoresis (PAGE). The pherogram shows the electrophoretic protein pattern of 12 horse blood sera after coomassie blue staining.

_Kärntner Blondvieh The Kärntner Blondvieh is an autochthonous, critically endangered Austrian cattle breed, found in Carinthia and Slowenia. The animals feature high fertility, robust health, longevity and good fattening capacity. Kärntner Blondvieh is famous for best meat quality.

_Lipizzan horse Lippizan horses are one of the oldest horse breeds in Europe. This elegant baroque horse is used as saddle and driving horse as well as for moderate agricultural work. As a Paneuropean horse breed, the Lipizzan horse is part of the European cultural heritage.

_Capillary electrophoresis After PCR amplification, the length of fluorescence labeled DNA fragments is automatically determined by laser detection during capillary electrophoresis. Data are analyzed by suitable software. In this system the PCR amplificates are automatically applied.

_European Lipizzan studs Until World War I Lipica was the Imperial Stud of the Austro-Hungarian Empire, where breeding Lipizzan horses is continued until today. However, the horses for the Spanish Riding School are provided from the Austrian Federal Stud Farm of Piber. Several other important Lipizzan stud farms exist in Europe: Topol'cianky, Szilvásvárad, Beclean, Fagaras, Djakovo and Monterotondo.

_The Vienna Spanish Riding School The exclusive aim of breeding Austrian Lipizzans is their use in the High Dressage School. The picture shows a Lipizzan School Stallion and the horseman, celebrating a trained exercise in the famous Vienna Spanish Riding School.

_In situ hybridisation Gene expression often anticipates developmental decisions. Here the expression of a mid/hindbrain boundary specific transcription factor (En2) is visualized by in situ hybridisation. At this stage the top view of a fish embryo (medaka) shows a uniform neural tube with eye vesicles forming anteriorly. Slightly later during embryogenesis morphological changes are initiated which execute the separation of midbrain and hindbrain.

_Medaka fish Female and male medaka (Japanese killifish). Living in rice fields these robust fish tolerate extreme conditions, an advantage for housing the animals in the laboratory.

_Zebrafish embryo Zebrafish embryo of one day. Full transparency and accessibility of the embryos are some of the advantages which recently made fish a valuable model organism for vertebrate development.

_Bernese Mountain dog The Bernese Mountain dog is a very popular breed. Nevertheless, the population is burdened with some genetic diseases. In a population genetic study we investigated the prevalence and heritability of the diseases to enable a breed specific breeding strategy.

_MRD The multifocal retinal dysplasy (MRD) is an autosomal recessive defect, occurring in numerous dog breeds. The ophtalmoscopic picture shows simple retinal folds in the tapetal fundus. Affected dogs have no impaired vision.

_Beagle Beagle hounds are high in huntsmen favour to course in packs or solitary on squirrels, rabbits and hares. Because of their small size and friendly character, beagles are popular companion dogs and the favorite laboratory dogs in medical research.

_Labrador Retriever Labrador Retriever  are  used as gun dogs as well as as pets. To help the breeders in producing healthy dogs, a study is conducted to find out prevalence and heritability of important diseases in the Labrador population and to define an effective breeding strategy to avoid genetic diseases in this breed.

_Puppy The choice of a puppy from a litter is a far-reaching decision for the dog keeper. The new companion for the next ten years should be healthy, free of genetic diseases,  friendly and bearer of all the desired characteristics which are specific for the breed.

_Dog litter A well performing and healthy litter, which is free of genetic diseases is not self-evident in several dog breeds. Breeders are recommended to let  their breeding animals be investigated for the breed specific diseases. Animals bearing breed specific diseases or genetic defects should be excluded from breeding.

_Siamese cat Coat pigmentation in siamese cats is caused by a temperature-sensitive tyrosinase.  If you cut the hair of a Siamese cat in a brighter region, it will re-grow dark because of the lower temperature of the hairless skin. Veterinarians should inform the owners of colourpoint cats about that fact before shaving  them in preparation for surgery.

_Tissue culture In vitro primary cell culture and permanent cell lines can be grown for different experiments. Cells, originating from various tissues and organisms can be cultivated. Tissue culture is a suitable system for tumor genetics, developmental biology and immunobiology. Aim of the in vitro tissue culture is the validation and/or replacement of in vivo experiments.

_Abnormal karyogram In the karyogram, metaphase chromosomes of a cell are identified and sorted by size, centromer position and banding pattern and are accordingly arranged in an allocation table. Homologous chromosomes are associated. In the picture we show an abnormal trypsin G-banded karyotype of a cat: Both A1-homologues are involved in a fusion event. In addition, the homologue of chromosome A2 is absent.

_FITC labeling The image shows FITC (Fluorescein-isothiocyanat) -labeled telomeric DNA on metaphase chromosomes of a cat lymphocyte. Counter-staining was done with propidium iodide.

_lFleckvieh 81,3% of the Austrian herd-book cattle belong to the Fleckvieh breed (simmental cattle). In Austrian the Fleckvieh is used for two purposes which is meat and milk production.

_Microscope Microscopes are indispensable tools in cytogenetics, tissue culture and animal reproduction. They are used to evaluate chromosomal preparations,  to visualize in vitro cultivated cells and to carry out oocyte and embryo manipulations.

_Hatching During their migration in the oviduct, embryos are spheric and protected by a glycoprotein-layer. In the uterus, hatching is the prerequisite to get in contact with the mucous membrane.

_Two cell stage embryo The picture shows a two cell stage embryo in mammalian development. The first cleavage of the bovine zygote takes place one day after fertilization.

_US guided investigation Ultrasound (US) guided systems can be used for (pregnancy)-diagnostic purposes, as well as for the recovery of oocytes or (amnionic)-fluids.

_Follicle punction Oocytes, used for in vitro production (IVP) of embryos, are collected from living donor animals. Oocytes are obtained by punction of follicles in different maturity stages or by oviduct irrigation under endoscopic control. Oocytes are picked up (OPU) and proceeded to in vitro maturation (IVM) and/or in vitro fertilisation (IVF) applications.

_Embryo In vitro cultivation In vitro production of embryos: Embryos are grown in vitro up to the blastocyst stage (7days). Then they are suitable for the bloodless transfer into the uterus of the recipient.

_Large White pigs Half of the Austrian herd-book pig breeds belong to the breed "Large White".  The animals feature high fertility, high number of raising piglets, rapid growth and minimum stress susceptibility. They are used for the production of crossbred sows in the hybrid breeding management.

_Rabbit embryo Rabbit embryos are surrounded by a thick mucin layer which is necessary to form a physiological placenta. The embryo in the picture is in the hatching state.

Rabbit stable Rabbits are held in small groups until they reach their sexual maturity. Then they are kept in separate cages as shown. For the housing of the animals constant light programs are used. Rabbits are mainly active at dawn and at dusk.

_Rabbit milking Transgenic animals are used for the production of recombinant proteins in the milk ("gene farming"). Milking rabbits is done by imitating the suckling of the puppies, while the doe is lying in a relaxed state.

_Milk Rabbit milk obtained with the milking machine is pipelined in a collecting pot.  A doe with a body weight of 4-5 kg can produce up to 1/4 liter milk per day. Rabbit milk features a very high fat content.

Laboratory rabbit We use rabbits with different genetic backgrounds. In the reproduction experiments hybrid rabbits bred for high quantity of meat are used. Selection of rabbits for high quantity of milk is still in progress.

_Micro-manipulator The micro-manipulation equipment is the essential tool to carry out microinjections. It consists of an inverse microscope and two micro-manipulators, the left one is carrying the retaining pipette, the right one is guiding the injection pipette. While the retaining pipette is holding the embryo in position by underinflation, DNA solution is injected into the pronuclei of the fixed zygotes.

_Cryoconservation Long-term storage of biological material is carried out in liquid nitrogene. The storage temperature is -196°C. If properly stored, semen, embryos and tissues can be kept for some decades without essential damage.

_PCR The polymerase chain reaction (PCR) is an enzymatic amplification process, where primer sequences are extended in the presence of template DNA during numerous amplification cycles. Reaction temperature is changing periodically for the denaturation of the DNA, for the annealing of the primers and for extension. Amplification of the defined DNA fragment during this process is approximately by the factor 106-108.