Method for embryo transfer in sheep
SUBSTANCE: the present innovation deals with individual matching donor sheep to recipient sheep at similar antigenic composition of blood types being correspondent to the value of antigenic similarity index being ra=0.51-1.00, where ra - antigenic similarity index. Moreover, the mentioned antigenic similarity index should be calculated by the following formula: where S - the number of similar antigens in a donor sheep and in a recipient sheep, n1 - the number of detected antigens in a donor sheep, n2 - the number of detected antigens in a recipient sheep. The present innovation enables to increase the level of adaptability of transferred ovine embryos by 25%.
EFFECT: higher efficiency of embryo transfer.
3 ex, 4 tbl
The technical field to which the invention relates.
The invention relates to the field of biotechnology of farm animals, in particular to embryo transfer in sheep.
The level of technology
Method of transplantation of embryos from cows less productive at the present time is one of the most common biotechnological techniques. This advanced technique has been successfully used in the intensification of reproduction of sheep. At the same time, a significant limiting factor is low (from 20 to 60%) accepted embryos after implantation in the body of the recipient.
Known methods and techniques to enhance the engraftment of early embryos in sheep during their transplant.
With the aim of enhancing the engraftment of transplanted embryos is recommended to enter at the top of the horns of the uterus of the recipient (Ernst L.K. Transplantation of embryos of farm animals / Lkist, Neergheen. - M.: Agropromizdat, 1989. - S-129).
It is also known that the successful settling down transplanted embryos in sheep depends on the stage of their development and synchronization of the estrous cycle in the recipient with the stage and the age of the embryo. Good results can be an exact match of the estrous cycle of the recipient with the age of the embryo (Beeham D., Sreenar J.M. Embrio transfer in farm animals Ed.K.J.Betteridge, Canada, Department of agricul. - 1977. - Monograph. 16. - P.16).
The best results engraftment get when transplanting recipients not 1, but several embryos (K. Kasymov embryo transfer in sheep / Kosimov, Galimzyanov, Nawano etc. // Zootechnics. - 1994. No. 8. - P.29-30). In addition, as a test of the high engraftment use a classification assessment of the suitability of embryos for transplantation (Sergeev, NI Bioengineering embryo transfer / Niedergeses, Sautepan // Sheep. - 1987. No. 2. - P.36-40).
There is a method of selecting horses that are resistant to grazing the clamp and its associated diseases, which includes the determination of the genetic parameters of blood, the genetic parameters of blood use polymorphic proteins and enzymes of the blood and the degree of heterozygosity, while sustainable animal is determined by the presence of phenotype serum arylesterase HB, level of heterozygosity 45-100% and the absence of transferrin phenotype FF (RF Patent No. 2212791, class. And 01 To 67/02, 12 Q 1/44).
The closest biotechnological solution to the proposed method (prototype), from among known methods for enhancing the engraftment of transplanted from donor to recipient embryos method is based on the definition of blood groups in cows-donors, recipients, bulls make the her and matching them so that to their antigenic spectrum (blood group) as little as possible was different from the antigenic spectrum of the embryo (Sukhova THAN the Accepted grafts cattle with regard to immune status manufacturers, donors, manufacturers / N.O. Sukhova, Wedplan, Vastaava etc. // Dokl. RAAS. - 1992. No..11-12. - P.27-30.; Verevochki PS Immunogenetics in breeding cattle / Psservice, Net - Kuibyshev, 1988. - 104 S.).
Significant disadvantages of the known methods is the low number obtained at the optimum stage of development of early and late blastocyst embryos due to the asynchrony of ovulation in the ovaries, the complexity of their vital evaluation, the wide variability in performance transplantation because of the complexity of influencing factors, the expenditure of a large number of embryos from high value animals to obtain acceptable results, which significantly reduces their effectiveness.
One of the reasons for the low engraftment of transplanted embryos may be immune and genetic incompatibility between donor and recipient. When embryo transfer is still unclear immunogenetic interaction system "embryo-animal-recipient", which, in some cases, successful implantation and conflict-free development of transplant in organisms is e priemysel mother, in other cases lead to its rejection. Identifying factors such mechanism could significantly improve the efficiency and effectiveness of the method embryo transfer.
Disclosure of inventions
The task of the invention is to develop a method that provides a higher level of engraftment of transplanted early embryos after their transplantation from sheep donor to the recipient.
The technical result that can be achieved using the proposed method, is to increase the level of engraftment of transplanted early embryos after their transplantation from sheep donor to the sheep-recipients.
The technical result is achieved by using method of embryo transfer in sheep, including individual selection of sheep donor to the sheep recipients, with individual selection carried out with similar antigenic composition of the blood groups, corresponding to the index value antigenic similarities from rand= from 0.51 to 1.00.
The method of selection of sheep-donor embryos to the sheep-recipients is as follows.
The selection of the elite herd of donor embryos and less productive sheep-recipients is carried out by any of known methods (instruction technology organizations on artificial insemination and transpl is ncacii embryos of farm animals. M., 2000. - 174 C.; Ernst L.K. Transplantation of embryos of farm animals / Lkist, Neergheen. - M.: Agropromizdat, 1989. - S-129.).
Then they held immunogenetic determination of blood groups in the immunological reaction of hemolysis and agglutination using monospecific immunodiagnostics with the subsequent index calculation antigenic similarity (IAS, rand).
The index calculation antigenic similarities conducted according to the following formula:
where S is the number of similar antigens in sheep donor and sheep-recipient;
n1- the number of identified antigens in sheep donor;
n2- the number of identified antigens in sheep recipient.
Estimated index is close to zero, means the minimum genetic similarity, and, on the contrary, the index is close to unity indicates maximum similarity of genetic parameters of donors and recipients. Further manipulations to obtain and embryo transfer are carried out by a known method (K. Kasymov embryo transfer in sheep / Kosimov, Galimzyanov, Nawano etc. // Zootechnics. - 1994. No. 8. - P.29-30.), that is illustrated by the following examples.
In the breeding season were selected 6 sheep donor, characterized by elite productivity, and 36 sheep-recipients with lower productivity. Donors and p is cipients were pre-selected blood samples and defined antigens of erythrocytes. Based on the immunogenetic characteristics between them were calculated indexes antigenic similarity (IAS, rand) blood group.
Multiple ovulation in sheep donor induced by the introduction of 12-14 sexual cycle 24 mg follicle-stimulating hormone (FSH) followed by injection of 125 μg of prostaglandin PGF 2 alpha Estrofan". Synchronization of the ovulatory process in the ovaries was provided by intravenous 800 units of human chorionic gonadotropin (HCG). Embryos from donors was extracted on day 5 after insemination (day 0) and after quality assessment, were transplanted recipients. Selection of pairs "donor-recipient" carried out the largest index antigenic similarities. Thus, on the value of the IAS donor recipients were divided into 3 groups.
The implementation of the invention
Examples of a specific implementation method of embryo transfer in sheep.
Example 1. In the first group consisted of a pair of "donor-recipient"with low genetic similarity blood group (rand=0.1 to 0.5) (table 1).
Selection of pairs of donor-recipient
|Animals||Index antigenic similarities (rand)||Antigens|
The effectiveness and efficiency of transplantation was determined by the level of engraftment of embryos, by number of lambs received-grafts and immunogenetic data examination of authenticity of origin (table 3, 4).
Example 2. The second group (experiment 2) entered a couple with a similar range of blood groups (ra=0,1-1,00).
Selection of pairs of donor-recipient
|Animals||Index antigenic similarities(rand)||Antigens|
|randor =0.51 to 1.0|
|Retz is pient||0,75||AA||Ab||Bd||Be||Ca||Da||Ma|
The floor is built, the results are shown in table 4.
Example 3. In the third group (prototype) definition of IAS between donors and recipients and individual selection of pairs "donor-recipient" was not performed (table 4).
Conducted by comparing the immunogenetic characteristics of the triad: father-mother-donor-offspring-transplant" genetic analysis showed that all lambs received 1, 2, and 3 experimental groups were obtained from transplanted zygotes: blood transplants matched groups of sheep blood donor and sheep, semen which they were seeded. The antigens of the recipient, from which were obtained the lambs-grafts, in their blood were absent, as shown in table 3.
Genetic examination of the reliability of the origin of the lambs-grafts
|RAM-manufacturer||Ab, Bb, Bc, Bd, Bg|
|The ewe-donor||AA, Ab, Bc, Be, Bi, Ca, Da|
|The ewe-recipient||AA, Be, Bi, Ca, Da, Ma|
|Descendant-transplant||Ab, Bb, Bc, Bd, Bg,|
The results of biotechnological experiments conducted in triplicate, indicate quite clear dependence of the rate of engraftment of embryos at transplantation and from the code, the antigenic similarity of groups of blood donors and recipients (table 4).
The settling down of embryos in sheep after transplantation, depending on the antigenic index of similarity of donors and recipients
|The antigenic index of similarity (IAS), rand||Used recipients||Received lambs||The effectiveness of transplantation, %|
|0,10-0,50 (experiment 1)||12||24||13||7||6||54,2|
|0,51-1,00 (experiment 2)||12||24||21||11||10||87,5|
Therefore, the proposed method of selecting sheep donor and recipient largest index antigenic similarity (IAS) blood groups among them in the boundary values of ra=0,51-1,00 has a number of positive differences and significant advantages compared with prototype:
- improving embryo implantation by 25% in selecting the optimal combination of donor and recipient blood groups can significantly reduce the consumption of embryos and loss of embriodered;
- significantly (71,4%) increases the number of lambs-grafts, and therefore, the efficiency of embryo transfer as a biotechnological method.
Method of embryo transfer in sheep, including individual selection of sheep donor to the sheep recipients, characterized in that the individual selection of sheep donor to the sheep-recipients spend with similar antigenic composition of erythrocytes when the value of the index antigenic similarity rand=from 0.51 to 1.00, which is determined by the formula
where rand- the index of the antigenic similarity;
S is the number of similar antigens in sheep donor and sheep-recipient;
n1- the number of identified antigens in sheep donor;
n2- the number of identified antigens in sheep recipient.
FIELD: medical equipment.
SUBSTANCE: device can be used in microbiology. It characterizes by peculiarities of taking microorganisms from surface of nose polipi for direct electron microscope testing. Device has rod with cap. Rod is made of iron. Two-sided sticky carbonaceous band is fixed onto cap of rod. Top sticky surface of band is brought into contact with mucous of polipi of nose for getting preparation-print of micro content followed by viewing it in scanning electron microscope. Device doesn't cause traumatism of patient's mucous and allows determining L-shaped bacteria quickly as well as their sizes, shape and types of reproduction onto polyp tissue.
EFFECT: improved efficiency of detection.
3 dwg, 2 ex
SUBSTANCE: method involves carrying out clinical biological examination of a teenager. When finding sensory disorders manifested as distorted taste and smell perception related to iron deficiency in organism in patients periodically observed at the age of 10-15 years and/or at the examination time, risk of primary pathological attraction to psychoactive drugs is to be predicted.
EFFECT: high reliability of prognosis.
SUBSTANCE: method can be used for measuring concentration of glucose in sample's fluid as tissue fluid. For performing the method the measuring device is used which has working part of detector, second working part of detector and control part of detector. Fluid's sample is applied onto measuring unit and electric current being proportional to concentration of matter in sample's fluid is measured in any part of detector. Electric currents are compared for setting difference. If the difference exceeds preset threshold value, the index of error generates to show that volume of sample is not sufficient.
EFFECT: improved precision of measurement.
2 cl, 2 dwg
FIELD: medicine, morphology.
SUBSTANCE: one should treat dura mater (DM) anatomical object with 3%-hydrogen peroxide solution. Moreover, DM should be preimpregnated into aqueous Evans blue solution for about 2-5 min. Visualization of lymphatic vessel should be implemented according countering external surface of its wall and gas that filled the vessel. The innovation enables to increase significance in detecting DM lymphatic vessels due to accurate visualization of vascular lumen.
EFFECT: higher efficiency and accuracy of detection.
SUBSTANCE: method involves carrying out clinical diagnostic examination and taking material sample from boundary separating intact and tumor tissue zones. Morphometry is applied to blood vessels of 120-130 mc large diameters. Mean blood vessel wall thickness is determined. The value being equal to 20-30 mc, repeated remote gamma-therapy course is prescribed. The value being greater than 30 mc, repeated radiation therapy course is considered to be inexpedient.
EFFECT: differentiated approach to prescribing repeated radiation therapy course; avoided risk of complications.
FIELD: veterinary science.
SUBSTANCE: one should study blood samples in 1-4-mo-aged healthy animals by the main parameters, moreover, the values of analyzed blood values by Table 2 should be transformed into indices (xi) to fill in Table 3, then one should detect average arithmetic value Mi=∑xi/n, where n - the number of tested animals or analyzed parameters, and xi - index value and according to Mi value one should evaluate the level of natural resistance in one animal or in group of animals, moreover, index 0.4 corresponds to low resistance level, 0.6 - to unsatisfactory level, 0.8 - to satisfactory one and 1.0 - to high resistance level. As seven main blood parameters one should analyze the following ones: the number of erythrocytes and leukocytes, percentage of neutrophils and lymphocytes in leukocytic formula, leukocytic index, the content of hemoglobin and that of total protein.
EFFECT: higher accuracy of evaluation.
1 cl, 1 dwg, 3 tbl
FIELD: medicine, stomatology.
SUBSTANCE: the suggested method deals with detecting deviation of biophysical value against the standard that should be calculated by the difference of polarized and nonpolarized fluorescence of buccal epithelial cells. The innovation provides broadened diagnostic possibilities.
EFFECT: higher accuracy of diagnostics.
FIELD: engineering of devices for analyzing multi-component dispersion environments, possible use for express-analysis of presence of given object in biological substance of complicated composition.
SUBSTANCE: database is composed of non-resilient dissipation spectrums and distribution of spectral characteristics of electromagnetic radiation intensiveness in zone of minimal values of probe radiation dissipation diagram, consisting of basic biological environment and researched object. Using aforementioned database is possible for controlling presence of viruses, bacteria, toxins (alcohol, drugs) in the organism, or determining foreign components (for example, admixture of soy albumen in milk, and also presence of heavy metal salts in there: lead, cadmium, zinc, mercury, arsenic and the like). Device has block with at least one source of coherent polarized monochromatic electromagnetic radiation, working cell for researched object, block for recording and reading optical information, a set of database cells for storing information about spectrums of combination dissipation or distribution of spectral characteristics of intensiveness of electromagnetic radiation in zone of minimal values of radiation dissipation diagram, and also control block, having output to personal computer. Block for recording and reading optical information has optical disk with recording layer on phase transitions, or holographic device or crystal, containing two layers with different crystalline grids and different conductivity types. Device can also include block with three radiation source in optical and/or adjacent infrared ranges, light splitter, spectrum analyzer for controlling spectrum particles and rotating device and is made with possible adjustment of angle between sources and receivers of radiation. Radiation sources can be built into single whole block with programmable central microprocessor unit, or programmable logical integral circuit.
EFFECT: increased efficiency.
4 cl, 2 dwg
FIELD: experimental medicine.
SUBSTANCE: the present innovation deals with preparing hepatic preparations for histological testing. In the course of an experiment on inducing hepatic pathology animals should be perfused with glutaraldehyde formaldehyde fixing agent through portal vein under conditions of narcosis. Then comes perfusion with incubation medium containing 0.2%-digitonin in Flickinger's solution at pH being 7.2 at 20-22°C. After incubation hepatic tissue should be reduced to be placed into 0.2%-digitonin for 4-10 h, and then samples should be washed with 0.1 M cacodylate buffer and postfixed with 1%-solution upon cacodylate buffer for 2 h. Then it is necessary to conduct dehydration to be put into araldite followed by contrasting with uranylacetate and lead citrate. The method enables to increase degree of contrast due to fixing hepatic tissues in situ and optimal matching incubation medium.
EFFECT: higher efficiency.
FIELD: medicine, oncology.
SUBSTANCE: in plasma of peripheral blood in children one should detect cathepsin D activity and antiproteolytic activity and calculate coefficient of their ratio. At obtaining the value of coefficient being equal 10.5 ± 2 one should detect lymphadenopathy of benign genesis in a child to choose a waiting-type tactics at prescribing antiphlogistic therapy. At coefficient value ranged 30-100 on should detect malignant lymphoproliferative process in a child to immediately fulfill an excision biopsy of a lymph node along histological studying and at proving the diagnosis special therapy should be indicated. Application of the present method enables to conduct additional objective differential-diagnostic test under disputable cases at the stage being before carrying out histological analysis of lymph nodes and prescribe necessary therapy for every concrete case in due time.
EFFECT: higher accuracy of differential diagnostics.
3 ex, 1 tbl
SUBSTANCE: method involves using product containing spermatozoa treated by drying with freezing to humidity level of 1% and having injured membrane or spermatozoon head so that spermatozoon nucleus retains its genetic validity enough for fertilization. The spermatozoon heads fertilize an isolated oocyte after rehydration and microinjection being done. The retained genetic integrity is enough for fertilizing an oocyte and producing living descendants. Method involves collecting living mature spermatozoa, making spermatozoa suspension in special purpose physiological medium, freezing the spermatozoa suspension for producing frozen spermatozoa, drying the frozen spermatozoa or spermatozoa heads in vacuum to humidity level of 1%, making rehydration of spermatozoa or spermatozoa heads with injured membrane and selecting those retaining nucleus of genetic integrity. The selected spermatozoa or spermatozoa heads are used for fertilizing isolated oocytes with living descendants being produced.
EFFECT: enhanced effectiveness in producing living descendants.
39 cl, 5 dwg, 1 tbl
SUBSTANCE: one should affect 1-14-d-aged chickens with "MINI-EXPERT-T" apparatus at 7.5-13.5 Hz frequency according to "Alpha-rhythms" program per 20 min thrice daily at 4-h-long interval, then one should affect 15-42-d-aged chickens with "Transfer-P" apparatus for energy-informational transfer of medicinal preparations by applying either Monotard MS or Protophan N.M. The innovation increases poultry safety and body weight gain in poultry.
EFFECT: higher efficiency.
2 ex, 7 tbl