149 results about "Embryo transplantation" patented technology

Disclosed is a method for using a first polar body and a second polar body as well as a single-cell embryo to carry out whole-genome non-exponential amplification and high-throughput genome sequencing, so as to perform preimplantation genetic diagnosis for genetic disease and testing for pathogenic genes causing repeated miscarriages. The method of the present invention comprises the following steps: (1) obtaining oocytes and embryos, and carrying out separation and genome amplification of first and second polar bodies and single-cell embryos; (2) establishing a genome sequencing library and sequencing, and carrying out bioinformatic analysis of the genome to obtain a gene spectrum and information concerning the number of copies of chromosomes and fragments thereof; (3) determining the chromosome ploidy of the polar bodies and embryos as well as information concerning defects, replication and point mutation in the chromosome fragments; (4) selecting normal or suitable embryos for implantation.

The invention provides an external fertilization and embryo transplantation information checking method. The method comprises the following steps that (1) identity information corresponding to a sperm, an ovum, an embryo and a matrix is stored in a database; (2) relevant identity information of culture dishes which carry the sperm, the ovum and the embryo and relevant identity information of a matrix identity recognition ring are recognized through the RF technology; (3) searching is performed on the database to compare whether the obtained identity information is matched with the database or not, and pairing is correct if yes; if not, pairing is wrong. According to the external fertilization and embryo transplantation information checking method, the defects in the prior art are overcome; under the condition of no manual intervention, the identity information of the culture dishes which carry the sperm, the ovum and the embryo and the identity information of the matrix identity recognition ring are recognized through the RF technology during external fertilization and embryo transplantation, and the recognized identity information is compared. In this way, the accuracy of the external fertilization and embryo transplantation can be guaranteed.

The invention discloses a preparation method for a myostatin knock-out pig, belonging to the technical field of biology. The preparation method is as follows: generating virus particles capable of expressing shRNA of myostatin gene of a targeted pig in a 293Ft cell; then infecting the fibroblasts of a piglet by using the produced slow virus; wherein after 48 hours, the expression of Myostatin genes in the fibroblasts is inhibited by fluorescence quantitative polymerase chain reaction (PCR) detection. In the invention, the slow virus particles can be used for carrying out microinjection on perivitelline space of a fertilized egg of the pig, can lead the virus nucleic acid to be integrated to an embryo gene group so as to express the shRNA, and can produce a transgenic pig with the myostatin of being knocked out after embryo transplantation. The slow virus particles also can be used for producing the transgenic pig by using a sperm vector method. By utilizing the preparation method, the cumbersome operation of knocking out the homologous targeting gene in the traditional method can be avoided, the virus consumption is less and the stable passage can be achieved.

The invention relates to sheep X/Y sperm-separated frozen semen, the production method and application thereof. The sheep X/Y sperm-separated frozen semen provided by the invention contains a frozen balance solution A, a frozen balance solution B including 12 percent glycerin cryoprotectant and a sheep X or Y sperm-separated frozen semen. The invention applies the difference of DNA content of sheep X and Y sperms and separates X and Y sperms through flow cytometry so as to manufacture the sex-control frozen semen and the frozen storage condition is 20 to 30min treatment under -90 to 120 DEG C. The frozen semen provided by the invention controls the lamb sex through artificial insemination or embryo transplantation. With high separation efficiency and sex control accuracy up to 90 percent, the frozen semen is suitable for industrialization production and culture, thus greatly increasing the culturing effect.

The invention discloses a protein extraction and treatment process at different time points (including early, middle and late periods of secretion: LH_2, LH_5, LH_7, LH_9, LH_10 and LH_12, and LH_0 on the day of injection of hCG) in a human endometrial tissue secretion period, and a protein spectrum and a phosphorylation modification mass spectrum are carried out to obtain molecular expression modes at different time points in the secretion period. The molecular expression modes can provide a basis for further knowing the dynamic change of the human endometrial tissue secretion period, and is beneficial to judging an endometrial implantation window period and guiding the embryo transplantation time in an assisted reproduction technology.

The invention discloses an equus oocyte vitrification cryopreservation fluid, a cryopreservation method and an application. The equus oocyte vitrification cryopreservation fluid comprises a base solution (BS), an equilibrium solution (ES) and a vitrification solution (VS), wherein the VS is composed of the following components by volume percent: 10-20% of glycol (EG), 6-10% of dimethyl sulfoxide (DMSO), 6-10% of propylene glycol (PROH), 15-25% of inactivated fetal calf serum (NCS), 8-15% of carboxylate E-polylysine (COOL-PLL) and the balance of an HEPES solution. Each liter of the VS includes 0.3-0.8mol of saccharose. The invention has the beneficial effects that the vitrification cryopreservation fluid has low cytotoxicity, high permeability and obvious vitrification effect, meanwhile can obviously reduce the generation of ice crystals in freezing and re-warming processes, can protect cells from physical injury, can increase the survival rate of oocyte, is suitable for the vitrification cryopreservation of equus oocytes or embryos of horses, donkeys and the like, and is beneficial to the popularization and application of artificial insemination and embryo transplantation technology.

The invention relates to the field of biotechnology, in particular to a method for constructing a lethal gene systemic knockout mouse model with a CRISPR/Cas9 system. The method includes the followingsteps that 1, an sgRNA sequence for efficiently identifying a lethal gene PAM region after knockout is designed; 2, mRNA or protein of Cas9 is mixed with the sgRNA designed in step 1, microinjectionis performed on any blastomere cell in a mouse two-cell embryo with the mixture, embryo transplantation is performed after injection, and strain gene identification is performed to obtain a chimeric positive founder with the lethal gene knocked out; 3, the positive founder and a wild-type mouse are mated to generate an F1 generation, a heterozygous F1-generation mouse with the lethal gene systemically knocked out is finally obtained after gene identification, and construction of the mouse model capable of realizing passage propagation is completed. Compared with the prior art in which a lethalgene systemic knockout mouse model is constructed by ES cell gene targeting, the method in the technical scheme has the advantages that the operation steps are simple, the operation difficulty is low, the production cycle is short, and the cycle is only about 4 months.

The invention discloses embryo secreting type endogenous polypeptide PDBCM and application thereof. The amino acid sequence of the embryo secreting type endogenous polypeptide PDBCM is as shown in SEQ ID NO.1. The polypeptide PDBCM can enhance BIRC6 expression, reduce apoptosis proteins Smac and reduce caspase-9 expression, thus promoting formation of blastula. The embryo secreting type endogenous polypeptide PDBCM can be applied to preparation of a medicine capable of promoting the formation of the blastula and/or improving the quality of the blastula or preparation of blastula culture liquid in an in vitro fertilization-embryo transplantation process.

The invention relates to a system for detecting ectopic pregnancy. The system comprises: a data acquisition module used for acquiring data of a 14th-day hCG level (hCG14) after embryo transplantationof a subject and a 21st-day hCG level (hCG21) after embryo transplantation of the subject, and acquiring data of main infertility factors of the subject; a data processing module used for further processing the data acquired in the data acquisition module; and a module used for calculating the probability of ectopic pregnancy by utilizing the data processed in the data processing module so as to calculate the probability of ectopic pregnancy of the subject after in-vitro insemination embryo transplantation or clear ovulation period natural pregnancy, and determining the probability grouping ofectopic pregnancy of the subject based on a calculation result.

The invention discloses an optimized mice somatic cell nuclear transplantation method. According to the mice somatic cell nuclear transplantation method, three kinds of cells, including cumulus cell, fetal fibroblasts, and embryonic stem cell, are adopted to construct reconstructed embryos; the activation efficiencies of three reconstructed embryos in different activation mediums are compared, development efficiencies in different solutions are compared, and pregnancy efficiencies of different embryo transplantation methods are compared; it is concluded that calcium-free KSOM activation medium is a universal high efficiency activation medium, and the activation efficiency is about 93.5%; KSOM-AA culture medium is suitable for in vitro culture of the reconstructed embryos of cumulus cells and embryonic stem cells, and the best culture medium for fetal fibroblasts is alpha MEM culture medium; when the reconstructed embryos are at 2-cell period, nuclear transplanted animals can be obtained via transplantation of embryos through bilateral fallopian tubes, wherein for each fallopian tube, transplantation of 15 embryos is carried out. The optimized mice somatic cell nuclear transplantation method is capable of increasing reconstructed embryo cleavage rate, blastula development rate, cloning animal birth rate, and survival rate, so that the success rate of obtaining embryonic stem cells from the reconstructed embryos is increased.

The invention provides a construction method of an SDK2 gene mutation mice model. The mice model comprises the following steps of designing and constructing sgRNA capable of specifically identifying an SDK2 gene on the basis of a CRISPR/Cas9 system, injecting the sgRNA and a targeting vector constructed on the basis of the sgRNA into a mice fertilized egg; and after embryo transplantation, screening out F0-generation mice with SDK2 gene mutation from the output mice, and hybridizing the F0-generation mice with wild type mice to obtain an F1-generation mice model with SDK2 gene mutation. The mice model constructed by the method can be stably passaged, the action mechanism of the SDK2 gene in mice hereditary cataract can be conveniently researched in practical application, and under the condition that human patient research materials are not easy to obtain and are restricted by medical ethics, the SDK2 gene can be stably cloned. The mice model provided by the invention can become an important tool in the research of hereditary cataract, and a research model capable of realizing stable heredity is provided in the research of pathogenesis, treatment methods, drug screening, cataract surgery and the like.

The invention discloses a reconstructed oocyte of a deaf model pig and a construction method of the reconstructed oocyte, and also discloses the deaf model pig and a construction method and application of the deaf model pig. According to the reconstructed oocyte of the deaf model pig, the construction method of the reconstructed oocyte and the construction method of the deaf model pig, by knockingout a pig OSBPL2 gene, the reconstructed oocyte of a deaf mini-pig is obtained, and in combination with a CRISPR/Cas9 gene editing technology, a somatic cell nuclear transfer technology and an embryotransfer technology, high efficiency and feasibility of the construction method for the genetically engineered deaf model pig are proven. Through the construction method of the deaf model pig, genotypes and deafness phenotypes of human OSBPL2 gene mutations can be precisely duplicated, the pig of which an OSBPL2 gene is knocked out has typical characteristics of human autosomal dominant hereditary hearing loss (DFNA67), therefore, a reliable big animal model can be provided for disease research of the human hereditary hearing loss, and the deaf model pig can be applied to inner-ear gene repairing, inner-ear hair cell regeneration, hearing reconstruction and the like.

The invention discloses a sperm competition fertilization dish and an application method thereof. The sperm competition fertilization dish comprises a dish body and a dish cover for enclosing an opening of the dish body. A plurality separator plates arranged in the dish body divide the dish cavity into an ovum accommodation zone and a sperm accommodation zone respectively at two sides of the dish body, and Z shape long channel communicating the ovum accommodation zone and the sperm accommodation zone. The invention simulates sperm dissociation path in human natural fertilization, realizes optimization of sperms, effectively guarantees that only high-quality sperms with enough vitality can arrive the and combine with the ovum, thereby raising growth rate of fertilized embryos and success rate of embryo transplantation, and finally guaranteeing excellent quality of the progeny.

The invention discloses a cutting operation liquid for equine animal embryos, and a preparation method and application thereof. The cutting operation liquid includes a sodium lactate mixed solution, penicillin, streptomycin, and one, two or three selected from bovine serum albumin, calf serum and inactivated equine serum. The preparation method includes the steps of: placing the sodium lactate mixed solution in a thermostat at 37 DEG C for 0.5-6 h for standby; and adding penicillin, streptomycin and one, two or three selected from bovine serum albumin, calf serum and inactivated equine serum into the sodium lactate mixed solution in accordance with the component contents, so as to obtain the cutting operation liquid for embryos. The cutting operation liquid is suitable for non-surgical method for embryo transplantation of horses, cattle and donkeys. The invention has the beneficial effect of convenience for material acquisition, low price, simple preparation and convenience for usage, is very suitable for scale production of non-surgical method for embryo transplantation of equine animals, and is favorable for popularization and application of embryo transplantation technology.

The invention discloses a breeding method for transgenic pigs expressing sIFITM3 (swine interferon induced transmembrane 3) genes. The breeding method comprises the steps as follows: firstly, the construction of swine fibroblast cell lines stably expressing the sIFITM3 genes comprises the following steps: the step a refers to the construction and the linearization of pcDNACAsIITM3 vectors, wherein, primers are designed, lymph node tissue RNA (Ribose Nucleic Acid) of pigs is extracted, the sIFITM3 genes are obtained through RT-PCR (Reverse Transcription-Polymerase Chain Reaction) amplification, ethanol precipitation is carried out, and sterile water is used for dissolving; the step b refers to liposome transfection, wherein, swine fibroblasts expressing sIFITM3 are constructed; the step c refers to screening and proliferation of nuclear donor cells transfected with the sIFITM3 genes; secondly, recombination blastula acquisition based on a manual nucleus transplantation method comprises the following steps: the step a refers to the preparation of recipient cells; the step b refers to enucleation and injection of the nuclear donor cells; and thirdly, the embryo transplantation and the acquisition of the transgenic pigs comprise the following steps: the step a refers to embryo transplantation, wherein, bred blastulas are transferred to a fallopian tube of a surrogate sow, and then detection is carried out; and the step b refers to transgenic individual identification. The breeding method is feasible, and is simple and convenient to operate; in addition, the pigs are the transgenic pigs expressing sIFITM3, and have the potential capability to resist animal virus such as foot-and-mouth disease virus, Japanese encephalitis virus, swine influenza virus and the like.

The invention provides a culturing method of a new strain of mutton goats with black heads and white bodies. The method is characterized in that a Boer goat is regarded as a male parent, a Macheng black goat is regarded as a female parent, and through a BLUP method, a marker-assisted selection (MAS) technology and combination of a conventional culturing method, by means of crossing fixup and embryo transplantation and propagation, the new strain of mutton goats with a high reproductive rate, a good meat performance and high temperature and humidity resistance are selected finally. According to the culturing method, the Macheng black goat is regarded as the female parent, the Boer goat is regarded as the male parent, and through the BLUP method, a multiple genes pyramiding technology and the combination of the conventional culturing method, the new strain of mutton goats with hair colors, high reproductive rate, meat performance and high temperature and humidity resistance is cultured; the characteristic of the black heads and white bodies is stable, the growth and meat performances of the new strain of the mutton goats are much higher than those of the Macheng black goat, the growth performance, the crude feed resistance, the disease resistance and the stress tolerance are higher than those of the Boer goat, and the production performance level of modern mutton goat breeds is achieved.

Compositions and methods for determining endometrial receptivity to embryo implantation and for detecting and treating castration-recurrent prostate cancer are provided. The methods comprise measuring the level of melanoma antigen gene protein-11 (MAGE-11, also referred to as MAGE-Al 1) in an endometrial or prostate tissue sample. The level of MAGE-11 protein or mRNA can be correlated to endometrial receptivity to embryo implantation in a female human or nonhuman primate, or to the presence of castration-recurrent prostate cancer in a male patient in need thereof. Methods are described whereby MAGE-11 may serve as a target for vaccine development in the treatment of castration-recurrent prostate cancer. Methods for monitoring endometrial maturation, for diagnosing infertility, and for in vitro fertilization in a female human or nonhuman primate are also provided. Compositions of the invention include antibodies that specifically bind MAGE-11 and oligonucleotide primers useful for detecting MAGE-11 mRNA, as well as kits containing such antibodies or primers.

The invention discloses a cultivation method of beef cattle hybridized by Charolais cattle, Simmental cattle and local cattle. The method includes: selecting a Tongjiang cow as the female parent; performing superovulation on the Tongjiang cow; selecting Simmental breeding bull sperm to perform artificial insemination; collecting the embryo of the Tongjiang cow; selecting a Charolais surrogacy cowto perform embryo transplantation; detecting 2 genotypes related to the growth traits of a first-filial-generation cow; selecting an F1-generation cow as the female parent; performing superovulation on the F1-generation cow; selecting Simmental breeding bull sperm to perform artificial insemination; collecting the embryo of the F1-generation cow; selecting the first-filial-generation cow to perform embryo transplantation; detecting 2 genotypes related to the growth traits of a second-filial-generation individual; selecting the cattle, which is good in body shape and growth speed and has dominant genotypes, hybridized by Charolais cattle, Simmental cattle and local cattle, and using the artificial insemination, superovulation and embryo transplantation technologies to expand propagation. Bythe method, the workload of scientific and technical personnel can be lowered, and cultivation time can be shortened.