19 results about "Leucosis" patented technology

The application belongs to the technical field of biology and particularly relates to an ELISA kit for detecting chicken astrovirus type II antibodies and a preparation method. The Cell-ELISA experiment is carried out by using LMH cells infected with chicken astrovirus type II to detect the chicken astrovirus type II antibodies. The method has no cross reactivity with positive serum of Newcastle disease virus, avian influenza virus, chicken infectious anemia virus, avian leukosis virus, Marek's virus and chicken infectious bursal virus, which indicates that the method has good specificity. The Cell-ELISA method is used to detect the chicken serum sample to be detected by using whole virus infection, and is assembled into a kit for measuring chicken astrovirus antibodies in serum. The kit can be used for serological diagnosis of chicken astrovirus type II infection, monitoring of the level of antibodies, epidemiological investigation and the like, and provides a novel and effective detection means for prevention and treatment of chicken astrovirus type II.

The invention relates to a primer probe combination for identifying E, J and K subgroups of avian leukosis viruses, a detection method and application of the primer probe combination. The detection system for identifying the avian leukosis virus subgroup comprises an upstream primer ALV-E-F, a downstream primer ALV-E-R and a probe ALV-E-P of a gp85 gene conserved region of targeted ALV-E, and an upstream primer ALV-J-F, a downstream primer ALV-J-R and a probe ALV-J-P of an env gene conserved region of targeted ALV-J, the upstream primer ALV-K-F, the downstream primer ALV-K-R and the probe ALV-K-P are used for targeting a gp85 gene conserved region of ALV-K. The detection system for identifying the avian leukosis virus subgroups has no cross reaction with other ALV subgroups and common avian pathogens, and has strong specificity.

The application provides a composition, a method and an application for improving the titer of mouse leukemia virus, and belongs to the technical field of biotechnology.The method comprises the following steps: before a host cell is infected with xenotropic mouse leukemia virus, the host cell is pretreated by using SC79; during virus infection of the pretreated host cell and during cell passage, mifepristone is added to a culture medium; and during continuous virus harvesting, LR3 IGF-1 is added to the culture medium.Through adding additives with different functional mechanisms at different stages of virus production, the application realizes dynamic optimization of the whole process of virus production.The experimental results show that the combination of SC79, mifepristone and LR3 IGF-1 can increase the titer of xenotropic mouse leukemia virus by 14.32 times, which is significantly better than the effect of single additive or two additives in combination.The application solves the problems of low titer of virus stock solution and low production efficiency in the prior art.

The application discloses application of a LOXL2 gene in resisting avian leukemia virus infection and belongs to the technical field of biotechnology.The nucleotide sequence of the LOXL2 gene is shown as SEQ ID NO.1.The application provides a new gene target point LOXL2 related to resisting avian leukemia virus infection, and proves the role of the LOXL2 gene in regulating avian leukemia virus replication.Inhibition of LOXL2 gene expression in host cells can promote avian leukemia virus replication, and overexpression of the LOXL2 gene in host cells can inhibit avian leukemia virus replication.The application provides favorable theoretical guidance and technical support for application of the LOXL2 gene as a target point in developing drugs for treating avian leukemia, breeding transgenic animals resisting avian leukemia, and constructing an avian leukemia virus infection model, and has a wide application prospect and extremely high market value.

The invention provides a molecular diagnosis method for avian leukosis virus and Marek's disease virus mixed infection. The molecular diagnosis method comprises the following steps: (S01) collecting diseased material tissues; (S02) carrying out nucleic acid extraction on the sick material tissue of the sick chicken; (S03) detecting the nucleic acid by using specific primers of avian leukosis and Marek's disease; (S04) preparing a PCR (Polymerase Chain Reaction) detection system, and respectively adding ALV-env gene amplification primers and MDV-meq gene amplification primers for gene amplification; (S05) carrying out 1% agarose gel electrophoresis detection on the amplification product in the step (4), and carrying out sequence determination after determining a target band; (S06) carrying out BLAST comparison on the sequences obtained by sequencing, carrying out gene sequence analysis and drawing an evolutionary tree; and (S07) determining the virus type according to the homology and the evolutionary branches, so as to realize rapid diagnosis of ALV and MDV mixed infection, strain characteristic analysis and exclusion of other similar epidemic diseases, and provide technical support for accurate prevention and control of AL and MD in the poultry industry, provenance purification and epidemiological investigation.

Provided is a kit for rapid detection of avian leukosis virus subgroup J (ALV-J) based on a CRISPR / Cas13a system. The method is based on the combination of the CRISPR / Cas13a system and recombinase aided amplification (RAA) for ALV-J detection. An oligonucleotide probe is designed as a substrate for CRISPR / Cas13a trans-cleavage and produces a detectable signal. The method can substantially improve detection sensitivity by amplifying a detection signal twice by RAA and T7 transcription. The detection method further exhibits excellent specificity, allowing for clear differentiate from other avian viruses. It does not require expensive experimental equipment and special laboratory environment, and it is rapid and efficient. The method is of great significance for biological research and on-site detection of ALV-J.

The invention discloses siRNA (small interfering Ribonucleic Acid) for inhibiting the expression of a chicken RASGRP3 (Reduced Amplification of Sequence Growth Protein 3) gene and application of the siRNA, the siRNA comprises a positive-sense strand and an antisense strand, and is any one of RASGRP3-Anas-214, RASGRP3-Anas-846 and RASGRP3-Anas-1187. According to the specific siRNA aiming at the chicken RASGRP3 mRNA provided by the invention, in a chicken liver cancer cell line (LMH), after liposome-mediated transfection culture is performed for 48 hours, RT-qPCR is adopted to evaluate the transcript level, and a result shows that the relative expression quantity of the chicken RASGRP3 mRNA is remarkably reduced compared with that of negative control, and the repeatability is good. The polypeptide can be further used for inhibiting ALV-J replication, provides a new molecular intervention target for prevention and control of avian leukosis, and has important scientific research and potential economic values.

This invention discloses a mutated A subgroup A avian leukosis virus (ALV-A) receptor gene and its application in combating A subgroup A avian leukosis virus infection. This invention identified L55 and W69 as key functional amino acid sites on the receptor protein Tva, which mediates ALV-A infection of chicken cells. Mutation of these two sites resulted in ALV-A's inability to infect host cells. Furthermore, this invention discloses a method for constructing a DF-1 cell line resistant to ALV-A infection. This method utilizes CRISPR / Cas9 and flow cytometry screening to mutate the key amino acid sites L55 and W69 of the Tva gene as the ALV-A receptor. Results show that substitution sequences containing mutations at L55 and W69 can efficiently and site-specifically replace the corresponding sequences in the Tva gene; the Tva gene-edited cell line can resist ALV-A infection without affecting cell proliferation. This invention lays the foundation for further research on the function of the Tva gene and the establishment of new technologies for combating ALV-A infection.

The application discloses application of MARCH2 gene as a target point in inhibition of replication of avian leukosis virus (ALV), and further discloses a MARCH2 gene overexpression cell line, a construction method thereof and application of the MARCH2 gene overexpression cell line in inhibition of infection of different subgroups of ALV. The MARCH2 gene overexpression cell line is successfully constructed by a lentivirus packaging infection technology, and experimental results show that the cell line can significantly inhibit replication of different subgroups of ALV. The application proposes that overexpression of the MARCH2 gene or expression of a coded protein of the MARCH2 gene can effectively inhibit replication of different subgroups of ALV. The finding provides a technical means for future anti-disease breeding work against ALV infection by taking the MARCH2 gene as a target point, and has a wide application prospect.

The utility model discloses an avian leukosis P27 antigen emulsifying device which comprises a working table and an L-shaped plate connected to the top of the working table, the L-shaped plate is connected with a stirring mechanism used for stirring a solvent, the stirring mechanism comprises a motor and a stirring head, the motor is used for driving the stirring head to rotate, the motor is connected with a sealing mechanism used for sealing a container, and the stirring head is connected with the sealing mechanism. The cover sealing mechanism comprises a sealing disc, a plurality of fixing cylinders, a plurality of moving rods and a plurality of springs, one ends of the moving rods are inserted into the fixing cylinders, the other ends of the moving rods are connected with the sealing disc, the springs are used for driving the sealing disc to move in the direction away from the fixing cylinders, and the motor is connected with an electric telescopic rod used for driving the motor to ascend and descend. And the operating platform is connected with a limiting seat for supporting the container. According to the utility model, the stability of the container can be improved, the leakage of a solvent is effectively avoided, the pollution to the working platform is reduced, and meanwhile, the lifting of the stirring head can improve the uniformity of antigen emulsification.

The invention belongs to the technical field of biology, and discloses a recombinant turkey herpesvirus, and a tandem epitope expression cassette is inserted between UL45 and UL46 of the recombinant turkey herpesvirus; the tandem epitope expression cassette is used for expressing a plurality of B cell epitopes and a plurality of T cell epitopes. According to the recombinant turkey herpesvirus, tandem epitopes are inserted into an HVT (turkey herpesvirus) carrier, and through experimental verification, good cellular immune response can be induced and generated in 1-day-old chicks and 18-day-old chick embryos by immunizing the multi-epitope recombinant HVT vaccine for one time, and compared with inactivated vaccines needing two-time immunization, the recombinant turkey herpesvirus can provide earlier and better challenge protection effect. Meanwhile, the recombinant turkey herpesvirus is verified to be capable of performing intraembryonic vaccination and generating an obvious immune protection effect. Meanwhile, the invention further provides a preparation method and application of the recombinant turkey herpesvirus.

The invention discloses a construction method and application of a feline leukemia virus mutant strain recombinant C subtype strain, and belongs to the technical field of virus gene engineering. The construction method comprises the following steps: designing a PCR primer containing specific amino acid mutation, and obtaining a fragment containing an RGD oligopeptide gene sequence through fusion PCR; amplifying a full-length gene sequence of the capsid protein p27 by taking the fragment as a template; the recombinant vector pFELVCRGD is inserted into a pTVT-FELVC vector to construct a recombinant vector pFELVCRGD; and transfecting the recombinant plasmid into a CD8T cell capable of stably expressing T7 RNA polymerase to rescue and obtain a mutant strain. The FELVCRGD strain obtained by the invention can be stably proliferated and subcultured in a cat lymphocyte FL74, and has a good immune protection effect on cats after being inactivated; the defects that existing FELVC in-vitro culture is difficult, traditional vaccine protection is insufficient and the like are overcome, and important technical support is provided for cat leukemia prevention and control and related basic research.

The application provides a composition, a method and an application for improving the titer of mouse leukemia virus, and belongs to the technical field of biotechnology.The method comprises the following steps: before a host cell is infected with xenotropic mouse leukemia virus, the host cell is pretreated by using SC79; during virus infection of the pretreated host cell and during cell passage, mifepristone is added to a culture medium; and during continuous virus harvesting, LR3 IGF-1 is added to the culture medium.Through adding additives with different functional mechanisms at different stages of virus production, the application realizes dynamic optimization of the whole process of virus production.The experimental results show that the combination of SC79, mifepristone and LR3 IGF-1 can increase the titer of xenotropic mouse leukemia virus by 14.32 times, which is significantly better than the effect of single additive or two additives in combination.The application solves the problems of low titer of virus stock solution and low production efficiency in the prior art.

The invention discloses an avian leukosis virus subgroup rapid detection kit and a detection method thereof, and belongs to the technical field of animal inspection and quarantine. Aiming at the defects of the existing detection technology, a conserved sequence of an env gene shared by A, B and K subgroups and a conserved sequence of a specific env gene of a J subgroup are selected as target regions, specific primer probes are designed, a reaction system and an amplification program are optimized, and the kit is assembled. The kit adopts a fluorescent RT-PCR (Reverse Transcription-Polymerase Chain Reaction) method, can simultaneously detect A, B, J and K subgroups and accurately distinguish endogenous and exogenous viruses without cross interference. The kit has the characteristics of high detection speed, high sensitivity, strong specificity, simplicity and convenience in operation and the like, is suitable for detecting various samples such as chicken plasma, seminal fluid and cloaca swab, can improve the breeding chicken purification efficiency, shorten the period and reduce the economic loss of the poultry industry, and has remarkable application value.

The application discloses an avian leukosis virus fluorescent quantitative PCR detection kit based on a pol gene and application thereof. The application is based on a fluorescent quantitative PCR technology, and specific primers and a TaqMan probe aiming at a pol gene conservative region of avian leukosis virus are designed to realize specific amplification and detection of virus nucleic acid. The core improvement of the application is that after nucleic acid is extracted from animal tissue, plasma and other samples, a DNA enzyme is added to perform digestion treatment, and residual host genome DNA in the sample is specifically degraded, so that a false positive problem caused by amplification of an endogenous pol gene is completely eliminated, and the specificity and accuracy of detection are obviously improved.

The invention discloses a BLV p24 recombinant antigen protein for detecting a bovine leukemia antibody, a test strip and application of the BLV p24 recombinant antigen protein. The amino acid sequence of the BLV p24 recombinant antigen protein is as shown in SEQ ID NO. 2. The eukaryotic recombinant p24 protein is prepared by adopting a baculovirus expression system, and the recombinant p24 protein expressed by the system can be correctly folded and modified after translation, better simulates native conformation and has higher sensitivity. And the His tag on the recombinant protein is beneficial to one-step purification to achieve higher purity, and better activity can be shown without enzyme digestion. The formed colloidal gold test strip is applied to detection of bovine leukemia, and the accuracy, the sensitivity and the detection are well improved.

The application relates to the technical field of antibodies, in particular to an antibody or antigen binding fragment specifically binding to p27 protein of avian leukosis virus and application thereof. The antibody or antigen binding fragment contains a heavy chain variable region and a light chain variable region, the amino acid sequence of the heavy chain variable region is shown as SEQ.ID NO.1 or SEQ.ID NO.5, and the amino acid sequence of the light chain variable region is shown as SEQ.ID NO.3 or SEQ.ID NO.7. The antibody or antigen binding fragment is positive to avian leukosis virus, which indicates that the antibody or antigen binding fragment has good reaction characteristics with the avian leukosis virus.

The application discloses a mutated K subgroup avian leukosis virus (ALV-K) receptor gene (Tva) and application thereof in resisting K subgroup avian leukosis virus infection. The application identifies key functional amino acid sites E53, L55, H59 and G70 of a receptor protein Tva mediating ALV-K infection of chicken cells, and finds that after the four sites are mutated, ALV-K cannot infect host cells. Further, the application also discloses a method for constructing a DF-1 cell line resisting ALV-K infection, which utilizes a CRISPR / Cas9 method and flow cytometry screening, and mutates the key amino acid sites E53, L55, H59 and G70 of the Tva gene as the ALV-K receptor. Results show that a replacement sequence containing the above site mutation can efficiently replace the corresponding sequence of the Tva gene; the Tva gene edited cell line can resist ALV-K infection without affecting the cell proliferation level. The application lays a foundation for further studying the function of the Tva gene and establishing a new technology for resisting ALV-K infection.

The invention belongs to the technical field of biology, and discloses a primer group and application thereof, and a kit for detecting RPA-CRISPR / Cas12a of ALV-F. According to the technical scheme, the primer group for specifically amplifying an ALV-F sequence is characterized by comprising an upstream primer and a downstream primer, the nucleotide sequence of the upstream primer is as shown in SEQ ID NO.1, and the nucleotide sequence of the downstream primer is as shown in SEQ ID NO.2; the nucleotide sequence of the downstream primer is as shown in SEQ ID NO. 2; based on a conserved region of an avian leukosis virus (ALV) F subgroup genome, a set of RPA primer group and crRNA with high specificity and high amplification efficiency are autonomously designed and verified. According to the design, the genetic diversity of target viruses is effectively covered, high specificity of detection on ALV-F subgroups is ensured, and the risk of cross reaction with other poultry pathogens is reduced to the maximum extent.