48 results about "Porcine tissue" patented technology

The invention discloses a porcine pseudorabies virus strain as well as an inactivated vaccine and applications thereof, belonging to the field of separation and application of the porcine pseudorabies virus strain. The invention firstly provides a porcine pseudorabies virus BJ strain separated from diseased pig tissues, and the microbial preservation number of the porcine pseudorabies virus BJ strain is CGMCC (China General Microbiological Culture Collection Center) No.7351. The invention discloses a method for preparing the inactivated vaccine by applying the porcine pseudorabies virus BJ strain. The method comprises the steps of culturing a virus strain to obtain a virus solution; adding an inactivator, and inactivating and concentrating the virus solution; and evenly mixing an adjuvant and the virus solution, and emulsifying to obtain the inactivated vaccine. The technological parameters of the inactivated vaccine preparation method are further optimized, and the immune protection efficacy and safety of the inactivated vaccine can be improved. Shown by the immune protection efficacy and safety tests, the porcine pseudorabies inactivated vaccine prepared has good immune protection efficacy and safety, and can be clinically used for preventing or treating porcine pseudorabies.

A method for separation the collagen from the various animal tissues is disclosed for preparing collagen solution and product using the same. The porcine tissues are processed to have proper form and size for acid-treatment. The acid-treatment is repeated with pepsin to separate type I or II collagens. The separated collagen is salt-treated for fractionation and ethanol-treated for obtaining 5˜10% of collagen from the initial tissue weight. The prepared tissues are processed for separating collagen through the collagen separating process. The separated collagen is processed for preparing product. The method for preparing product is comprised: treating a collagen solution having a predetermined concentration under a neutral condition at a low temperature, followed by overnight treatment at a temperature of 30 to 35° C.; concentrating collagen by centrifugation; and dissolving the thus-concentrated collagen in refrigerated weakly-acidic solvent or phosphate buffered saline (PBS), thereby preparing collagen having a concentration of 1 to 5 mg / mL.

The present invention relates to a method for enzymaticly digesting porcine fat, wherein fat cells are fragmented by using an ultrasonic wave-assisted enzymatic digestion method. The present invention also provides a method that is based on the foregoing method and that is for further refinement of advanced edible lard and fat tissue protein hydrolysate. The method comprises: extracting lard in in a cryogenic condition; obtaining the advanced edible lard after refining the lard; and obtaining a sauce by performing homogenizing, flavouring, thermal reacting, filling, and sterilizing on the protein hydrolysate obtained during enzymatic extraction.

The invention discloses a PCR primer pair for identifying or assisting identification of an animal tissue / or an organ and application thereof. The primer pair provided by the invention is at least one from five PCR primer pairs for identifying or assisting identification of cattle, sheep, pig, chicken and duck tissue / or organ, wherein the PCR primer pair for identifying or assisting identification of pig tissue / or organ is always included. The five PCR primer pairs for identifying or assisting identification of cattle, sheep, pig, chicken and duck tissue / or organ respectively consist of two DNA single chains shown as a sequence 1 and a sequence 2, two DNA single chains shown as a sequence 3 and a sequence 4, two DNA single chains shown as a sequence 5 and a sequence 6, two DNA single chains shown as a sequence 7 and a sequence 8, and two DNA single chains shown as a sequence 9 and a sequence 10 in the sequence table. An annealing temperature of 62 DEG C is employed for the five primer pairs to complete all PCR identifications at once and at a same temperature; besides, the PCR primer pair has strong singularity and a short detection time.

A method for separation the collagen from the various animal tissues is disclosed for preparing collagen solution and product using the same. The porcine tissues are processed to have proper form and size for acid-treatment. The acid-treatment is repeated with pepsin to separate type I or II collagens. The separated collagen is salt-treated for fractionation and ethanol-treated for obtaining 5˜10% of collagen from the initial tissue weight. The prepared tissues are processed for separating collagen through the collagen separating process. The separated collagen is processed for preparing product. The method for preparing product is comprised: treating a collagen solution having a predetermined concentration under a neutral condition at a low temperature, followed by overnight treatment at a temperature of 30 to 35° C.; concentrating collagen by centrifugation; and dissolving the thus-concentrated collagen in refrigerated weakly-acidic solvent or phosphate buffered saline (PBS), thereby preparing collagen having a concentration of 1 to 5 mg / mL.

The invention provides a Seneca valley virus SVV-ZM-201801 and an application thereof. Separation is performed from pig tissues, and sub-culture and plaque purification are performed, so that the Seneca valley virus is obtained. The separated strain can be stably proliferated on sub-culture cells, typical cytopathic effects are formed, and the virus titer is as high as 108.5-1010.5TCID50 / mL. The separated strain does not have obvious pathogenicity on piggies. The Seneca valley virus separated strain has good proliferating properties, and is good in immunogenicity. Vaccines prepared from the separated strain can induce the piggies to generate high-level neutralizing antibodies, and powerful technical support is provided for effective prevention and control of the Seneca valley virus.

The invention relates to the field of detection of veterinary drug residues, in particular to a confirmatory analysis method for efficiently detecting piperazine residues in poultry and pig tissues (muscle, liver, kidney and sebum). In this method, tissue samples are extracted, purified and derivatized by accelerated solvent extraction, and then detected by gas chromatography-tandem mass spectrometry (GC / MS-MS). Compared with other detection methods in terms of recovery rate, precision and sensitivity, the present invention has the advantages of simplicity, speed, accuracy and sensitivity.

Porcine nucleic acid sequences flanking potentially infectious porcine endogenous retroviral (PERV) insertion sites have been identified and isolated. The unique flanking sequences include porcine nucleic acid sequences that flank the 3′ end and porcine nucleic acid sequences that flank the 5′ end of PERV insertion sites. The present invention provides compositions and methods for detecting presence of PERV in a sample, particularly those with infectious potential. In addition, the invention relates to breeding of pigs or selection of porcine tissue that is free of infectious PERV for use as a xenotransplant tissue.

The invention relates to medicinal kallikrein (KLK1) and a preparing method and application thereof. The kallikrein does not contain KLK1c. Tissue kallikrein from porcine pancreas is composed of three components with different glycosylation degrees. A protein electrophoresis analysis result is three stripes, named KLK1b, KLK1a and KLK1c respectively in a molecular weight descending order, wherein KLK1c has the lowest glycosylation degree and the lowest specific activity. Compared with existing tissue kallikrein from porcine pancreas available in the market, the tissue kallikrein with KLK1c removed has the advantages of being higher in purity and stability, higher in biological activity, good in pharmacodynamic effect and smaller in side effect. The invention also provides a method for purifying porcine tissue kallikrein. The problem that KLK1b, KLK1a and KLK1c are hard to separate and industrialization can not be achieved is solved.

The invention discloses a nucleotide sequence for detecting swine NLRP6 (nod-like receptor family pyrin domain-containing protein 6) and application of the nucleotide sequence. The nucleotide sequence for detecting the swine NLRP6 is shown as sequences SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3 in a sequence table. The sequences SEQ ID NO.1 and the SEQ ID NO.2 respectively represent sense primers and antisense primers for detecting the swine NLRP6, and the sequences SEQ ID NO.3 represent fluorescent probes for detecting the swine NLRP6. The invention further discloses a method for detecting the swine NLRP6 by the aid of the primers and the probes. The nucleotide sequence, the application and the method have the advantages that as shown by results, the method is good in specificity and high in sensitivity, expression of the swine NLRP6 in swine tissue and organ samples can be quickly and accurately detected, the swine NLRP6 can be monitored in real time when NLRP6 inflammasome is researched, inflammation occurrence procedures and development procedures can be timely understood and mastered, and the nucleotide sequence, the application and the method have important theoretical significance and practical significance.

The invention belongs to the field of biochemistry and molecular immunology, and relates to a porcine neuronal interleukin B polypeptide, and a preparation method and application of a polyclonal antibody thereof. The preparation method comprises the following steps: coupling C-terminal modified polypeptide of the porcine neuronal interleukin B polypeptide with a carrier protein keyhole limpet hemocyanin KLH to obtain an NMB modified polypeptide-KLH coupled protein; emulsifying the NMB modified polypeptide-KLH coupling protein, and immunizing New Zealand white rabbits; collecting and separatingto obtain serum containing a rabbit anti-pig NMB modified polypeptide antibody; and purifying the prepared NMB polyclonal antibody by adopting a Protein G purification column method. A polypeptide antigen and the polyclonal antibody have the characteristics of simple preparation method, low cost and high titer, and can specifically bind to an NMB protein in pig tissues.

The invention discloses a rapid test card for simultaneously detecting PEDV and TGEV, and preparation and application methods thereof. The rapid test card comprises a card shell and a test strip arranged in the card shell. The test strip comprises a PVC bottom plate, a sample pad, a combination pad, a nitrocellulose membrane and an absorption pad, the sample pad, wherein the combination pad, the nitrocellulose membrane and the absorption pad are sequentially connected and pasted to the PVC bottom plate; and a sample adding hole and a result observation window are formed in the card shell. An anti-PEDV specific monoclonal antibody and an anti-TGEV specific monoclonal antibody are fixed on the nitrocellulose membrane and are used for respectively forming a first detection line and a second detection line; and the anti-PEDV specific monoclonal antibody marked by colloidal gold and the anti-TGEV specific monoclonal antibody marked by colloidal gold are fixed on the combination pad. According to the present invention, with the rapid detection card, the PEDV and the TGEV in a pig tissue excrement sample can be simultaneously detected, the sample adding only needs one time, the reaction is performed under the unified condition, and the rapid detection card has advantages of rapidness, high accuracy, high sensitivity, simple operation and the like.

The invention provides an African swine fever rapid detection test strip. The African swine fever rapid detection test strip is composed of a test strip and a sample treatment liquid. On the other hand, the invention also provides a method for preparing the test strip. The method comprises the following steps of 1) preparing an anti-African swine fever virus P30 protein monoclonal antibody and ananti-African swine fever virus P72 protein monoclonal antibody, 2) preparing a latex microsphere pad and a sample pad, 3) preparing a nitrocellulose membrane detection line and a quality control line,4) preparing and sub-packaging a sample treatment solution, and 5) assembling test strips. The African swine fever rapid detection test strip provided by the invention is suitable for detection of African swine fever viruses in porcine serum, porcine whole blood and porcine tissues, is strong in specificity, high in sensitivity, good in stability and high in detection speed, can be used for earlyscreening of the African swine fever virus, and is particularly suitable for on-site African swine fever infection diagnosis, epidemiological investigation, pig international trade quarantine inspection and the like.

The invention discloses a porcine-derived osteopontin (OPN) monoclonal antibody, a hybridoma cell strain thereof, and applications of the monoclonal antibody and the hybridoma cell strain. The hybridoma cell strain secreting the porcine-derived OPN monoclonal antibody is classified and named as a hybridoma cell strain Cy2, and is preserved in China Center for Type Culture Collection (CCTCC) on Dec.19,2019, wherein the preservation address is Wuhan University, Wuhan, China, and the preservation number is CCTCC NO:C2019293.The porcine-derived OPN monoclonal antibody is secreted by the hybridomacell strain or a passage cell strain of the hybridoma cell strain. The monoclonal antibody is suitable for rapidly and accurately detecting the content of OPN protein in pig tissues, semen or pig oocytes, so that the monoclonal antibody can be used for preparing related detection kits or colloidal gold test paper, and a firm foundation is laid for deep research of porcine-derived OPN.

Porcine nucleic acid sequences flanking potentially infectious porcine endogenous retroviral (PERV) insertion sites have been identified and isolated. The unique flanking sequences include porcine nucleic acid sequences that flank the 3' end and porcine nucleic acid sequences that flank the 5' end of PERV insertion sites. The present invention provides compositions and methods for detecting presence of PERV in a sample, particularly those with infectious potential. In addition, the invention relates to breeding of pigs or selection of porcine tissue that is free of infectious PERV for use as a xenotransplant tissue.

The present application relates to the field of veterinary diagnostic technology, and specifically to a probe composition, a primer composition, and a triple fluorescence PCR kit and related methods for differential diagnosis of three genes of different strains of African swine fever composed of the above-mentioned probe composition and primer composition. The kit includes primer probes specific to the B646L gene, the CD2v gene, and the I177L gene. The present application designs fluorescent PCR primers and probes for the B646L gene, the CD2v gene, and the I177L gene, respectively, and establishes a triple fluorescence PCR detection method, which can quickly identify African swine fever virus strains and gene-deficient strains. At the same time, it has no cross-reaction with swine fever virus, porcine reproductive and respiratory syndrome virus, pseudorabies virus, porcine epidemic diarrhea virus, porcine circovirus, porcine parvovirus, healthy whole blood, oropharyngeal swabs, nasal swabs, anal swabs, environmental swabs, pig tissues, etc., and has good specificity.

The invention relates to a method for detecting pig fat deposition by using phylaxin expression level, which comprises the following steps: 1) extracting total DNA from detected pig tissue, carrying out inverse transcription to obtain a cDNA sample; 2) carrying out PCR amplification on the cDNA sample from the step 1) by primer pair shown in SEQ ID NO:2 and 3 to obtain a gene fragment shown in SEQ ID NO:1; 3) carrying out PCR amplification on the housekeeping gene by primer pair shown in SEQ ID NO:5 and 6 to obtain a gene fragment shown in SEQ ID NO:4; 4) carrying out agarose gel electrophoresis on the PCR products from the step 2) and step 3), comparing the optical density value in a gel imaging system. According to the invention, the fat pigs and lean type pigs with 1 month age, 3 months age, 5 months age and 7 months age can be taken as detection objects, the expression level of fat cytokine phylaxin (resistin) can be detected, the protein level of resistin is taken for reflecting the deposition condition of the pig fat. The method of the invention has the advantages of rapidity, convenience, accuracy and no influence on normal growth of pigs, and is suitable for dynamically monitoring the fat deposition condition of pigs in the culture process.

The invention provides a triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, a kit and application, and relates to the technical field of biological detection. According to the triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, primers and probes are synthesized according to conserved sequences of porcine circovirus type 2, porcine circovirus type 3 and porcine circovirus type 4 in Genbank, and meanwhile, the kit is further constructed and used for porcine epidemic disease detection. 83 porcine tissue samples from Zhejiang Province are detected by utilizing an established fluorescent quantitative PCR system, and a result shows that the primer group is good in specificity, repeatability and stability, relatively high in consistency and relatively high in accuracy. According to the triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, the lowest detection values of PCV2 and PCV3 are both 10 copies / microliter, the lowest detection value of PCV4 is 1 copy / microliter, and the defects of low sensitivity, prone laboratory pollution and the like of the current triple PCR detection method for the porcine circovirus type 2, the porcine circovirus type 3 and the porcine circovirus type 4 are overcome.

The invention discloses a pig UBC promoter and application thereof, and belongs to the field of gene engineering. The pig endogenous UBC promoter is separated and identified, it is determined through cell tests that the pig endogenous UBC promoter can drive an exogenous gene to be efficiently, stably and widely expressed in pig tissue cells, and the problems that in the transgenic pig product preparation process, the exogenous promoter possibly faces gene silencing, expression inconsistency and low expression efficiency are solved. The porcine endogenous UBC promoter has practical application value in preparation of transgenic pigs.

The invention discloses a PCR primer pair for identifying or assisting identification of an animal tissue / or an organ and application thereof. The primer pair provided by the invention is at least one from five PCR primer pairs for identifying or assisting identification of cattle, sheep, pig, chicken and duck tissue / or organ, wherein the PCR primer pair for identifying or assisting identification of pig tissue / or organ is always included. The five PCR primer pairs for identifying or assisting identification of cattle, sheep, pig, chicken and duck tissue / or organ respectively consist of two DNA single chains shown as a sequence 1 and a sequence 2, two DNA single chains shown as a sequence 3 and a sequence 4, two DNA single chains shown as a sequence 5 and a sequence 6, two DNA single chains shown as a sequence 7 and a sequence 8, and two DNA single chains shown as a sequence 9 and a sequence 10 in the sequence table. An annealing temperature of 62 DEG C is employed for the five primer pairs to complete all PCR identifications at once and at a same temperature; besides, the PCR primer pair has strong singularity and a short detection time.

The invention discloses a pig IGFN1 protein polyclonal antibody, further discloses antigenic polypeptide for inducing the pig IGFN1 protein polyclonal antibody and belongs to the field of animal gene engineering. The antigenic polypeptide is formed by the amino acid sequence as shown by SEQ ID No: 1. The nucleotide sequence shown by SEQ ID No: 2 forms the antigenic polypeptide encoding genes. The pig IGFN1 protein polyclonal antibody is high in pig IGFN1 protein expression level detection specificity, and an important tool is provided for expression analysis and correlational studies of IGFN1 in pig tissues. The antigen sequence is long, and the detection sensitivity is high.

The invention provides a pretreatment kit for detecting ochratoxin A in pig tissue. The pretreatment kit is characterized by comprising a buffer solution, an extracting solution 1, an extracting solution 2, an IAC column, a leacheate and an elution solution. The invention also provides a method for detecting the ochratoxin A in the pig tissue by utilizing the pretreatment kit. The method is characterized in that after the pig tissue is pretreated by utilizing the kit, the ochratoxin A is quantitatively detected by utilizing liquid chromatography-mass spectrometry under a certain liquid and mass spectrometry condition. The kit and the method have characteristics of low detection limit and high sensitivity and are relatively high in stability, precision and accuracy.

The invention discloses a method for detecting a menbutone residue in pig tissue. The method comprises the steps as follows: firstly, the pig tissue subjected to intramuscular menbutone injection is weighed, cut up and then subjected to homogenate, and the tissue subjected to homogenate is extracted by using acetonitrile, then after centrifugation, the extracted liquid is subjected to rotary evaporation in vacuum with the temperature of 60 DEG C to dry, and after cooling, moving phase is added into the residue for ultraphonic dissolution and then centrifugation, and a liquid supernatant is collected and filtered by a millipore filter membrane with filtering particle diameter of 0.22 mu m, and the filter liquor is used as a test solution; secondly, a menbutone reference substance is weighed precisely, and after dissolved by acetonitrile and diluted by moving phase, the reference substance has a concentration of 9 to 11 mu g / mL, and a reference substance solution is obtained; thirdly, HPLC (High-Performance Liquid Chromatography) is adopted for testing the test solution and the reference substance solution, the chromatogram is recorded, and the menbutone residue in the tissue is calculated by peak area according to external standard method. The method is good in specificity, high in sensitivity and good in repeatability, can quickly detect the menbutone residue in the pig tissue, and is a reliable method for controlling the quantity of the menbutone residue in animal-origin food.

The present invention provides a Porcine parvovirus (PPV) and its application. The Porcine parvovirus of the present invention is isolated from diseased pig tissues, named PPV-ZM2021, and the preservation number in the General Microbiology Center of the China Microbial Culture Collection Management Committee is CGMCC No. 45271. This strain can stably proliferate on passage cells and produce typical cytopathic effects. The gene of the Porcine parvovirus strain of the present invention has special mutation sites and has excellent immunogenicity; it can reach a high titer through suspension culture; the vaccine prepared with this isolate can induce pigs to produce high levels of neutralizing antibodies and has good cross-protection; at the same time, the ELISA kit prepared with this isolate can better detect Porcine parvovirus antibodies in serum.

The invention discloses a polyclonal antibody of porcine CENP-A protein as well as a preparation method and application of the polyclonal antibody, and relates to the technical field of bioengineering. The preparation method comprises the following steps: immunizing an animal by using a pig CENP-A protein, and collecting and separating to obtain serum; performing extraction and purification treatment on the serum to obtain the polyclonal antibody; the amino acid sequence of the pig CENP-A protein is as shown in SEQ ID NO. 1. The polyclonal antibody of the CENP-A protein is relatively good in specificity and relatively high in titer, and the polyclonal antibody of the CENP-A protein, provided by the invention, can be used for specifically recognizing endogenous CENP-A protein in pig tissues or pig small intestine epithelial cell lines in immunoblotting analysis, and can be applied to a centromere region sequence combined with CENP-A captured by ChIP-seq.

The invention discloses a method for detecting the concentration of flurbiprofen in pig tissue, and belongs to the technical field of biological medicine detection. The method provided by the invention comprises the following steps: cutting a pig tissue sample treated by flurbiprofen plaster into pieces, adding a PBS (Phosphate Buffer Solution), and grinding by using a biological homogenizer to obtain homogenate; performing vortex treatment on the homogenate to obtain vortex homogenate, then mixing the vortex homogenate with an internal standard working solution, performing vortex treatment and centrifugation to obtain a supernatant, then adding a formic acid aqueous solution, performing vortex uniform mixing, and then performing analysis by using liquid chromatography-tandem mass spectrometry. The method provided by the invention has the advantages of good repeatability, wide linear range, high sensitivity, high recovery rate, simple operation and efficient analysis.

The invention discloses a liquid chromatography-tandem mass spectrometry determination method for altrenogest in pig tissue, which comprises the following steps: sample pretreatment: enzymolysis treatment: weighing 1.0 g of pig tissue sample which is pig liver, pig muscle or pig skin adipose tissue; a protease K solution is added into the pig tissue sample, pig liver or pig muscle tissue is subjected to enzymolysis for 1 h in a dark water bath at the temperature of 37 DEG C, and pig skin adipose tissue is subjected to enzymolysis for 30 min in a dark water bath at the temperature of 50 DEG C; performing ultrasonic treatment for 2 minutes after enzymolysis, and cooling to room temperature; according to the method, the purification efficiency and the specificity are greatly improved, and the problem of matrix interference is effectively solved. The MIP-MNPs prepared by taking altrenogest as a template molecule can specifically adsorb altrenogest and metabolites thereof through spatial structure matching and functional group action, so that non-specific interference of protein, fat and steroid impurities in pig tissues is eliminated, and the ionization inhibition effect of a matrix on mass spectrometry detection is remarkably reduced.

The invention discloses a freeze-dried microchip, a kit and a method for detecting African swine fever virus in pig tissue and pig blood feed products. The freeze-dried microchip is characterized in that a fluorescent PCR reaction system for detecting the African swine fever virus is fixed on the microchip through freeze-drying; the kit comprises a freeze-dried microchip, a bottle of mineral oil,a tube of positive control, a tube of negative control, a tube of diluent and a tube of nuclease-free water. A primer probe group is used for identifying the African swine fever (ASF) virus, and detecting whether to-be-detected pig tissue and a pig blood feed product sample contain the African swine fever virus or not. By utilizing the freeze-dried microchip, the kit and the method provided by theinvention, ASF virus is detected from pig tissue and pig-derived feed for the first time, it is proved that the freeze-dried microchip has an important application value on prevention and control ofthe ASF, is beneficial to finding and cutting off a virus transmission chain, and can effectively prevent large-scale outbreak.

The invention discloses a porcine OPN monoclonal antibody, its hybridoma cell line and application. The hybridoma cell line secreting porcine OPN monoclonal antibody, which is classified as hybridoma cell line Cy2, was deposited in the China Center for Type Culture Collection (CCTCC) on December 19, 2019, and the preservation address is Wuhan, China .Wuhan University, the preservation number is CCTCC NO: C2019293. The porcine OPN monoclonal antibody is secreted and produced by the above-mentioned hybridoma cell line or its passaged cell line. The monoclonal antibody is suitable for quickly and accurately detecting the content of OPN protein in porcine tissue, semen or porcine oocytes. Therefore, the monoclonal antibody can be used to prepare related detection kits or colloidal gold test paper, etc. The in-depth research of OPN has laid a firm foundation.