35 results about "Bactrian camel" patented technology

The invention discloses a natural nanometer antibody library for Bactrian camel phage display. The natural nanometer antibody library is prepared from the following steps: sampling Bactrian camel blood and spleen which are not immunized with any antigen, extracting total mRNA, reversely transcribing into cDNA, and amplifying VHH by nested PCR (Polymerase Chain Reaction); carrying out enzyme digestion on a pMECS phage display vector and the VHH by restrictive incision enzymes Pst I and Not I and connecting two fragments; electrically transforming a connected product into a competent cell TG1. The invention further discloses a construction method and application thereof in the sieving of PCT and NAGL nanometer antibodies. The constructed natural phage display library can obtain the nanometer antibodies with specificity and a detection function through sieving, and can solve the problem that as a camel cannot be immunized due to antigen factors, the corresponding phage display library cannot be constructed, and accordingly, the nanometer antibodies cannot be obtained. Meanwhile, the library can also solve the problem that the immunization of the camel spends long time.

The invention discloses a bactrian camel heavy-chain (HC) variable-domain antibody resisting a porcine circovirus 2 as well as a preparation method and an application thereof, and belongs to the field of a biotechnology. A PCV2 (Porcine Circovirus Virus) inactivated vaccine immunized bactrian camel is firstly utilized and a phage display technology is utilized to establish a PCV2 immune VHH antibody base; and a high-specificity variable domain antibody of heavy chains of HCAbs (VHH) which has a good affinity with the PCV, particularly a PCV2 virus antigen; and the antibody disclosed by the invention has an amino acid sequence shown as SEQ (sequence) ID NO.1 or an amino acid sequence which is shown as SEQ ID NO.2 and has the sequence isogeny being more than 95%. The PCV2 heavy-chain variable-domain antibody disclosed by the invention has the very important meanings on researching porcine circovirus, particularly researching a porcine circovirus 2-type infection mechanism and establishing rapid antigen detection with high sensitivity and specificity or diagnosing a reagent.

The invention discloses a nano antibody corresponding to apolipoprotein ApoE, and also discloses a gene sequence for coding the nano antibody and a host cell capable of expressing the nano antibody. By virtue of the nano antibody gene sequence and the host cell disclosed by the invention, the efficient expression of the nano antibody can be achieved in escherichia coli; and the nano antibody is applicable to the research and development of ApoE molecular detection reagents.

The invention discloses a bactrian camel VHH (variable domain of the heavy chain of HACbs) heavy-chain antibody for resisting foot-and-mouth disease AsiaI type viruses, a preparation method and use thereof, belonging to the field of biotechnology. In the method, recombinant AsiaI type virus-like particle antigens are used for immunizing bactrian camels; an AsiaI VHH immune antibody library is established with phage display technology; high-specificity single-domain antibodies having good affinity with foot-and-mouth disease AsiaI type virus antigens are screened; the antibody provided by the invention has amino acid sequence shown by SEQ ID NO.1 or amino acid sequence which has sequence homology more than 98% with the sequence shown by SEQ ID NO.1. The AsiaI VHH heavy-chain antibody provided by the invention has great importance for researching AsiaI type virus infection mechanisms and producing high-sensitive and specific antigen fast detection or diagnosis reagents.

The invention discloses a nanobody capable of resisting H9N2 subtype avian influenza viruses, a preparation method and an application. The amino acid sequence of the nanobody is as shown in SEQID NO:1. The nucleocapsid protein of the H9N2 subtype avian influenza viruses is expressed through a prokaryotic expression technique, then the nucleocapsid protein is used as an immunogen for immunizing Bactrian camels, a phage library is constructed, then a nanobody capable of resisting H9N2 nucleocapsid protein is obtained through screening by a phage display technique, then the nanobody and horse radish peroxidase (HRP) are subjected to fusion expression, and a fusion protein of the nanobody and the HRP is successfully prepared. When the fusion protein is applied to detection of antibodies capable of resisting H9N2 subtype avian influenza viruses (AIV) in chicken serum, the inventor finds that a detection method constructed through the fusion protein is high in sensitivity, and has the advantages that the operation is simple, secondary antibodies do not need to be used, and time consumed for detecting samples is short.

The invention discloses a porcine reproductive and respiratory syndrome virus Nsp2 protein nano-antibody and an application thereof. The nano-antibody is named Nb12, and the amino acid sequence of thenano-antibody is represented by SEQ ID NO.3. A camel is immunized with a truncated expression Nsp2 recombinant protein, camel peripheral blood lymphocytes are separated, and an immunized bactrian camel VHH gene is amplified by using RT-PCR, the amplified gene is connected into a pCANTAB 5E phage vector to construct a bactrian camel heavy chain antibody variable region library, 44 Nsp2 specific nano-antibodies are screened out through three rounds of panning by utilizing a phage display technology, tests prove that the nano-antibodies can specifically react with a Nsp2 protein, and the Nb12 has strong binding force. Antiviral test results show that when the virus inoculation amount is 0.01 MOI, the nano-antibody Nb12 shows a virus replication inhibition effect on both PAM cells and Marc-145 cells. Therefore, a new technical means is provided for subsequent research of the PRRSV Nsp2 and prevention, control and diagnosis of the PRRSV.

The invention relates to the technical field of genetic engineering and discloses a bactrian camel alpha-lactalbumin gene, a recombinant protein and a cloning method thereof. The gene has the characteristics of easy digestion and absorption, has a gene which is closely related to the functions of sterilizing and inducing apoptosis of tumor cells and has a nucleotide sequence from 4-432nd locus of the nucleotide in SEQ ID NO.1. The bactrian camel alpha-lactalbumin is one of main components of whey protein, has the function of regulating galactopoiesis, has the cytolysis activity, has the multiple functions of inducing cell growth inhibition and cell apoptosis and is an important protein in camel's milk. The established bactrian camel alpha-lactalbumin sequence and an evolutionary tree thereof can intuitively prove the difference between the bactrian camel alpha-lactalbumin and the protein sequence of other species, and experimental data are provided for addition of the camel's milk in food and further biochemical research; and therefore, the bactrian camel alpha-lactalbumin gene has high application value.

The invention discloses a nucleotide sequence for coding a Bactrian camel cytochrome P450, a Bactrian camel cytochrome P450 and application thereof, relating to the technical field of gene engineering. The main technical scheme is as follows: the nucleotide sequence is 29th-1580th sites in SEQ ID NO:1 in the sequence table; or the nucleotide sequence is hybridized with the 29th-1580th nucleotide sequence in SEQ ID NO:1 in the sequence table under highly strict conditions and used for coding the Bactrian camel cytochrome P450; or the nucleotide sequence has more than 90% of homology with the 29th-1580th nucleotide sequence in SEQ ID NO:1 in the sequence table and used for coding the Bactrian camel cytochrome P450. The Bactrian camel cytochrome P450 is coated by the nucleotide sequence.

The invention discloses an immunological kit for detecting florfenicol. A florfenicol immunodetection antibody is prepared by immunizing a Camelidae animal with a conjugate of a florfenicol derivativeand a carrier protein as a florfenicol complete antigen. The high-affinity florfenicol Camelidae animal antibody is obtained in the invention, the alpaca polyclonal antibody titer of florfenicol is 1:32000, and the Bactrian camel polyclonal antibody titer of florfenicol is 1:256000; and a florfenicol enzyme-linked immunoassay kit can be developed in the invention, the florfenicol detection limitof the kit is 0.3 ng / mL, the half-inhibitory concentration is 13.3 ng / mL, and the kit can achieve high-flux, fast and highly-sensitive florfenicol detection requirements. The kit can be used for the residual detection of florfenicol in livestock and poultry, aquatic products and other animal foods, has the advantages of easiness in operation, high in sensitivity and low price, is very suitable forscreening a large number of samples, and has wide application prospects.

The invention discloses bactrian camel source C-strain E2 VHH and application. The bactrian camel source C-strain E2 VHH and the application aim at solving the problem that production and preparation of high-purity high-appetency classical swine fever virus antibodies can not be obtained in the existing classical swine fever virus antigen diagnostic reagent research and development process. The bactrian camel source C-strain E2 VHH are characterized by being provided with a nucleotide sequence: SEQIDNO.1, SEQIDNO.2 or SEQIDNO.3. The invention further provides the application of the bactrian camel source C-strain E2 VHH in preparing a classical swine fever virus antigen testing diagnostic reagent. The bactrian camel source C-strain E2 VHH and the application have the advantages that the VHH coded sequence of C-strain antigen protein E2 is obtained for the first time through a Bactrian camel, ELISA experimental results show that the recombination VHH expressed in E.coli have the same antigen binding capacity as classical swine fever virus polyclone antiserum, and therefore the recombination VHH can replace the classical swine fever virus polyclone antiserum to be used for researching the classical swine fever virus antigen diagnostic reagent.

The invention relates to an optimal immune procedure and method of an immune camel. The immune method comprises the step of multipoint subcutaneous injecting at the back of a Bactrian camel, wherein the immune period is two weeks, the injected antigen dosage is 1mg / ml, and the immune is carried out three times. The invention further relates to a preparation flow and a method of a rabbit anti-camel IgG enzyme-labeled secondary antibody, a method for detecting camel anti-serum titer by applying the rabbit anti-camel IgG enzyme-labeled secondary antibody through ELISA and an optimal detection condition.

The invention relates to the technical field of genetic engineering and discloses a bactrian camel lysozyme protein gene, a recombinant protein and a cloning method thereof. The gene is a gene which is closely related to a gene related to glycoprotein synthesis of controlling nonheme iron associative property in the transferring family. The bactrian camel lysozyme protein gene and an evolutionary tree thereof can intuitively prove the difference between the bactrian camel lysozyme protein sequence and the protein sequence of other species, experimental data are provided for the functions of researching transfer of bactrian camel iron, realizing the broad-spectrum antibacterial and anti-virus function, regulating the in-vivo iron balance, promoting myelopoiesis, promoting cell growth, enhancing the body immunity and disease resistance and suppressing the tumor cells and disease treatment, a certain theoretical basis is provided for researching regulation of certain inflammatory responses and body immunity systems of the bactrian camel, other species and humans, and a certain biological basis is provided for research of body immune defense system; and therefore, the bactrian camel lysozyme protein gene has high application value.

The invention relates to the technical field of genetic engineering and discloses a bactrian camel H-FABP protein gene, a recombinant protein and a cloning method thereof. The H-FABP gene sequence of other species is referred, a cDNA sequence of the bactrian camel H-FABP protein gene is obtained through clone sequencing, the CDS sequence of the H-FABP gene of various species is subjected to homologous comparison, the structural features of the H-FABP gene are known and verified, and basic data are provided for a relationship for researching the H-FABP gene and the bactrian camel lipid metabolism. When H-FABP is combined with the traditional markers such as cTnI and CK-MB for detection, the H-FABP is detected in the early stage, and the cTnI is detected in the restoration stage. A patient suffering from cardiac event high risk is screened, the positive treatment strategy is conveniently taken, the expenditure of the patient and the lab resources can be saved, and reasonable and complementary monitoring of H-FABP and cTnI is an ideal choice of clinical treatment of ischemic heart disease.

The invention relates to the technical field of genetic engineering and discloses a bactrian camel A-FABP protein gene, a recombinant protein and a cloning method thereof. A nucleotide sequence of the bactrian camel A-FABP protein gene is obtained through clone sequencing, the CDS sequence of the A-FABP gene of various species is subjected to homologous comparison, the structural features of the A-FABP gene are known and verified, and basic data are provided for a relationship for researching the A-FABP gene and the bactrian camel lipid and energy metabolism. The bactrian camel A-FABP protein is a protein with multiple biological functions and has a key regulation function in energy metabolism and fatty acid metabolism; and therefore, due to the research of A-FABP from genes, proteins and the like, a novel hope is brought to prevention and treatment of obesity, insulin-risistant symdrome, type II diabetes and other diseases, and the bactrian camel A-FABP protein gene has high application value.

The invention relates to a marking method for indicating and distinguishing domestic and wild Bactrian camel DNA microsatellites in the technical field of bio-engineering. The steps of the method are as follows: ear tissue samples of Bactrian camels are collected, and genomic DNA is extracted according to the conventional phenol / chloroform method; PCR amplification is carried out on the genomic DNA of the Bactrian camels by means of using 16 pairs of designed primers; PCR products are separated by means of using non-denatured polyacrylamide gel electrophoresis, and genotype is classified according to the lengths of the PCR products; distribution of different microsatellite-marked genotypes in the Bactrian camels is compared, and the result shows that allele-specific genes of 3 microsatellite loci (VOLP-10, YWLL36 and YWLL38) exist between domestic and wild Bactrian camels; allele-specific amplified products are cloned and sequenced. The method can be operated simply and can distinguish the domestic Bactrian camels and the wild Bactrian camels at the genome molecular level, thereby providing an important means and method for researching the heredity and the evolution of the Bactrian camels, especially the identification and the protection of the wild Bactrian camels.

The invention provides a canine distemper virus H protein nano antibody and a preparation process thereof, and relates to the technical field of nano antibodies. The preparation process of the canine distemper virus H protein nano antibody comprises the following steps: S1, immunizing a bactrian camel with an inactivated vaccine; s2, separating cells; s3, amplifying a VHH gene and constructing a display vector; s4, constructing a nano antibody library and measuring the capacity of the library; s5, measuring the titer of the recombinant bacteriophage; s6, elutriating H protein specific recombinant bacteriophage; enriching the specific recombinant bacteriophage of the S7H protein; the method comprises the following steps: immunizing a camel with a canine distemper inactivated vaccine, separating peripheral blood lymphocytes of the camel, amplifying a VHH gene of a bactrian camel by using a nested PCR (Polymerase Chain Reaction) method, connecting the VHH gene into a pCANTAB 5E phage vector, constructing a phage display library, and screening by using a phage display technology to obtain three H protein specific nano antibodies, tests prove that the nano antibodies can specifically react with H protein, and have the characteristics of small molecular weight, good stability and high affinity.

The invention relates to the technical field of genetic engineering and discloses a bactrian camel pepsin A protein gene, a recombinant protein and a cloning method thereof. The bactrian camel pepsin A protein is a protein with multiple biological functions and has a key regulation function of regulating protein digestion; and therefore, due to the research of pepsin A from genes, proteins, enzymatic activity, inhibitors and the like, a novel enlightenment is brought to development of the fields of medicines and foods. The pepsin A gene sequence of other species is referred, a cDNA sequence of the bactrian camel pepsin A gene is obtained through clone sequencing, the CDS sequence of the pepsin A gene of various species is subjected to homologous comparison, the structural features of the pepsin A gene are known and verified, and basic data are provided for a relationship for researching the pepsin A gene and the bactrian camel digestion system.

The present invention provides a two-humped camel-based anti-goat pox virus VHH-4-1 and a use thereof, and belongs to the field of biotechnology. The anti-goat pox virus VHH-4-1 is prepared by VHH-4-1screening, expressing and purifying, functionalized labeling and activity verifying, anti-goat pox virus concentrating and purifying, optimizing various indicators of a kit, etc. Due to the fact thatthe provided kit uses the functionalizedly labeled VHH-4-1, the kit avoids steps of primary antibodies, secondary antibodies, anti-antibodies, etc., improves sensitivity and stability, shortens time,saves resources and breaks through drawbacks that traditional Elisa methods are time-consuming, laborious, etc. The provided anti-goat pox virus VHH-4-1 provides a basis for establishing sensitive goat pox virus clinical diagnosis method and studying a virus infection mechanism, and also provides a key marker for the developed goat pox virus antibody solid phase competition Elisa kit. In addition, goat pox attenuated vaccine is used to be subjected to cell enlarge cultivation, concentration and purification as a coating antigen to improves the sensitivity.

The invention discloses an immunological kit for detecting florfenicol. A florfenicol immunodetection antibody is prepared by immunizing a Camelidae animal with a conjugate of a florfenicol derivativeand a carrier protein as a florfenicol complete antigen. The high-affinity florfenicol Camelidae animal antibody is obtained in the invention, the alpaca polyclonal antibody titer of florfenicol is 1:32000, and the Bactrian camel polyclonal antibody titer of florfenicol is 1:256000; and a florfenicol enzyme-linked immunoassay kit can be developed in the invention, the florfenicol detection limitof the kit is 0.3 ng / mL, the half-inhibitory concentration is 13.3 ng / mL, and the kit can achieve high-flux, fast and highly-sensitive florfenicol detection requirements. The kit can be used for the residual detection of florfenicol in livestock and poultry, aquatic products and other animal foods, has the advantages of easiness in operation, high in sensitivity and low price, is very suitable forscreening a large number of samples, and has wide application prospects.

The invention relates to the technical field of genetic engineering and discloses a bactrian camel lysozyme protein gene, a recombinant protein and a cloning method thereof. The gene is a closely related gene which has the functions of resisting bacteria, diminishing inflammation and resisting viruses and can enhance the antibody activity and has a nucleotide sequence from 27-481st locus of the nucleotide in SEQ ID NO.1. The bactrian camel lysozyme sequence and an evolutionary tree thereof can intuitively show the difference between the bactrian camel lysozyme sequence and the protein sequence of other species, experimental data are provided for research on the hydrolysis of lysozyme of the bactrian camel and other species on the bacterial cell wall, a certain theoretical basis is provided for binding the lysozyme of the bactrian camel, other species and humans with viral protein with negative charges and inactivating the bound protein, and an important function of researching the functions of the lysozyme in human bacteria resistance, inflammation elimination and virus resistance; and therefore, the bactrian camel lysozyme protein gene has high application value.

The invention discloses a bactrian camel-based anti-goat pox virus VHH-4-2 and application thereof, and belongs to the field of biotechnology. The technical scheme includes: screening, expressing, purifying, functional labeling and activity verifying of anti-goat pox virus VHH-4-2, concentrating and purifying of goat pox virus, optimizing of various indicators of a kit, etc. The functionally labeled VHH-4-2 is used in the invention, thus avoiding operation steps of primary antibody, secondary antibody, anti-antibody, etc., improving sensitivity and stability, shortening time and saving resources. The bactrian camel-based anti-goat pox virus VHH-4-2 breaks through shortcomings that the traditional Elisa method is time-consuming, laborious, etc. The bactrian camel-based anti-goat pox virus VHH-4-2 lays a foundation for establishing a clinical diagnosis method for sensitive goat pox virus and for studying the infection mechanism of the virus; and the bactrian camel-based anti-goat pox virus VHH-4-2 also provides a key marker for the development of a goat pox virus antibody-immobilized competitive Elisa kit with complete proprietary intellectual property rights. In addition, the attenuated goat pox vaccine is used as a coating antigen after expanded culture, concentration and purification, so that the sensitivity is improved.

The invention relates to an Alxa bactrian camel superovulation method, and belongs to the technical field of animal production. According to the technical scheme, the implementation schedule, the medicine ingredients and the medicine dosage are determined, an Alxa bactrian camel superovulation procedure is set up, the blank that no bactrian camel superovulation procedure exists in China is filled in, estrus synchronization of animals is achieved, each experimental animal reacts to the superovulation procedure, and 7 to 22 functional corpora lutea can be formed. The medicine and the procedure are more suitable for bactrian camels and more economical compared with a foreign dromedary superovulation procedure.

The invention discloses a bactrian camel heavy-chain (HC) variable-domain antibody resisting a porcine circovirus 2 as well as a preparation method and an application thereof, and belongs to the field of a biotechnology. A PCV2 (Porcine Circovirus Virus) inactivated vaccine immunized bactrian camel is firstly utilized and a phage display technology is utilized to establish a PCV2 immune VHH antibody base; and a high-specificity variable domain antibody of heavy chains of HCAbs (VHH) which has a good affinity with the PCV, particularly a PCV2 virus antigen; and the antibody disclosed by the invention has an amino acid sequence shown as SEQ (sequence) ID NO.1 or an amino acid sequence which is shown as SEQ ID NO.2 and has the sequence isogeny being more than 95%. The PCV2 heavy-chain variable-domain antibody disclosed by the invention has the very important meanings on researching porcine circovirus, particularly researching a porcine circovirus 2-type infection mechanism and establishing rapid antigen detection with high sensitivity and specificity or diagnosing a reagent.

The invention discloses a bactrian camel VHH (variable domain of the heavy chain of HACbs) heavy-chain antibody for resisting foot-and-mouth disease AsiaI type viruses, a preparation method and use thereof, belonging to the field of biotechnology. In the method, recombinant AsiaI type virus-like particle antigens are used for immunizing bactrian camels; an AsiaI VHH immune antibody library is established with phage display technology; high-specificity single-domain antibodies having good affinity with foot-and-mouth disease AsiaI type virus antigens are screened; the antibody provided by the invention has amino acid sequence shown by SEQ ID NO.1 or amino acid sequence which has sequence homology more than 98% with the sequence shown by SEQ ID NO.1. The AsiaI VHH heavy-chain antibody provided by the invention has great importance for researching AsiaI type virus infection mechanisms and producing high-sensitive and specific antigen fast detection or diagnosis reagents.

The invention relates to a method for determining the titer of a swine transmissible gastroenteritis virus N protein antibody. The method comprises the following steps of: S1, performing an immune test on an experimental animal, comprising: determination of the titer of an N protein immune bactrian camel and an N protein antibody; S2, constructing a VHH bacteriophage library, comprising: amplification of a VHH gene, construction of a display vector, construction of a nano antibody library, and determination of the capacity of the library; and S3, elutriating an N specific nano antibody, comprising: library rescue and recombinant bacteriophage titer determination, N protein specific recombinant bacteriophage elutriation, protein specific recombinant bacteriophage enrichment and crude extract preparation, and ELISA detection of an N protein recombinant nano antibody.

The invention relates to a heat clearing and detoxifying Guangchang white lotus seed electuary and a production method thereof. Guangchang white lotus seed, deer gall, Loveyly Hemsleya Root, bactrian camel gallstone, Folium Polygoni Tinctrorii, seed of Japanese apricot, India Canna Rhizome, shrubalthea leaf, Hoya carnosa, honeysuckle flower and Indigofera trifoliate are adopted as main raw materials. The production method mainly comprises the steps of material selecting, drying, crushing, granulation and packaging to form a finished product. The heat clearing and detoxifying Guangchang white lotus seed electuary has the advantages of health, heat clearing and detoxifying effects, fast effectiveness, pure naturalness, and no toxic or side effects, has a good auxiliary treatment effect on main symptoms of wind heat colds, and is suitable for household eating.

The invention relates to a camel milk characteristic peptide fragment combination and an identification method, a specific characteristic peptide marker combination is found through detection by a high performance liquid chromatography-tandem mass spectrometry technology, the characteristic peptide fragment combination comprises characteristic peptide fragments for identifying authenticity of camel milk powder as shown in SEQ ID NO.1-SEQ ID NO.8, and single-peak and double-peak camel milk can be identified by SEQ ID NO.6-SEQ ID NO.8. Through a large number of experimental screening, the peptide fragment with stable property, high response and good repeatability is screened from the specific characteristic peptide fragments of species, the influence of enzymolysis property, protein modification and matrix effect on detection is eliminated, and the established method is high in sensitivity, good in accuracy and high in operability. Besides, the characteristic peptide fragment in the protein is selected as a detection object and is not influenced by protein stability and deep processing, detection of non-denatured and denatured proteins in a sample can be met, and the method is high in applicability. And an effective technical means is provided for camel dairy product adulteration monitoring and commodity label authenticity supervision.

The invention discloses a single-domain heavy chain antibody for detecting porcine epidemic diarrhea virus, a preparation method and application. The present invention adopts porcine epidemic diarrhea virus to immunize Alxa Bactrian camels, uses its peripheral blood lymphocytes to establish a single-domain heavy chain antibody library against porcine epidemic diarrhea virus, and couples the expressed and purified porcine epidemic diarrhea virus S protein to As an antigen on the enzyme plate plate, the single domain heavy chain antibody library was screened immunologically by using phage display technology, and the single domain heavy chain antibody gene against the S protein of porcine epidemic diarrhea virus was obtained, and the single domain heavy chain antibody gene was transferred into the large intestine In Bacillus, a single-domain heavy-chain antibody strain highly expressed in Escherichia coli was established, and the gene sequence of each single-domain heavy-chain antibody strain was analyzed to obtain the amino acid of the VHH chain of the single-domain heavy-chain antibody against the S protein of porcine epidemic diarrhea virus Sequence SEQ ID NO.8. The antibody of the invention has good stability and specificity, and can be used for detecting and treating PEDV.

The application provides a tamde virus, the virus strain is TAMV / XJ01 / Xinjiang / China / 2018, and the deposit number of the virus strain is CGMCC No.17585; the virus strain is from the tick subtype of Xinjiang Bactrian camel It is isolated from Hyalomma asiaticum; the application also provides primers and probes for detecting the virus, which have guiding significance for the mechanism of TAMV virus infection in ticks and the monitoring and control of the virus.