8 results about "Mdck cell" patented technology

This invention provides a method for constructing and applying MDCK cells stably integrated with disialotyltransferase, relating to the field of biotechnology. The construction method includes: infecting MDCK cells with lentiviral particles, followed by culture and screening to obtain MDCK cells stably expressing porcine ST3GAL1 and porcine ST6GAL1 genes; the lentiviral particles carry porcine ST3GAL1 and porcine ST6GAL1 genes; the nucleic acid sequence of the porcine ST3GAL1 gene is shown in SEQ ID NO.1; the nucleic acid sequence of the porcine ST6GAL1 gene is shown in SEQ ID NO.2. This construction method is simple and convenient, and the constructed MDCK cells can stably express porcine ST3GAL1 and porcine ST6GAL1 genes, effectively improving the culture efficiency of swine influenza virus, and thus can be used in the preparation of swine influenza virus.

Disclosed is an adapted Madin-Darby canine kidney cell line capable of suspension culture in the absence of serum, and a chemically-defined medium for culture of the adapted MDCK cell line. Further disclosed are culture methods for growing the adapted MDCK cell line and methods for producing a vaccine from the adapted MDCK cell line grown in the chemically-defined medium.

The invention provides application of levistilide A in preparation of anti-influenza virus drugs, and belongs to the technical field of biological medicines. The influenza virus is an H1N1 virus. In-vitro anti-influenza experiments find that levistilide A can significantly inhibit infection of H1N1 virus and significantly improve cytopathy caused by influenza virus infection. The H1N1 virus half inhibitory concentration (IC50) of the levistilide A in MDCK cells is 1.462 M, and the H1N1 virus half inhibitory concentration (IC50) of the levistilide A in A549 cells is 0.455 M. The invention proves that the levistilactone A has a remarkable anti-H1NA virus effect, can be used for preparing the anti-influenza virus medicine, and has an excellent application prospect.

Provided are an FLNB gene-knockout MDCK cell line, a construction method therefor and the use thereof. The FLNB gene-knockout cell line is named MDCK-FLNB-KO, and has been deposited in China Center for Type Culture Collection (CCTCC) on November 20, 2024, and the accession number is CCTCC NO: C2024391. The FLNB gene knockout uses CRISPR / Cas9 technology, wherein a sgRNA is designed for exon 1 of the FLNB gene of MDCK cells, an FLNB gene knockout vector plasmid 1 and plasmid 2 are constructed, and the plasmid 1 and the plasmid 2 are transfected into the MDCK cells to obtain the FLNB gene-knockout MDCK cell line. Compared with the original MDCK cell line, the obtained cell line has a lower adhesion ability and a higher yield when applied to influenza virus culture.

This invention discloses a method for efficiently isolating human parainfluenza virus based on suspended MDCK cells. By using the fully suspended passaged MDCK cell line for human parainfluenza virus isolation and culture and optimizing the inoculation process, single-generation virus amplification can be completed within 3 days, significantly shortening the cycle, improving virus isolation efficiency and success rate compared to traditional adherent culture methods, and enhancing experimental reproducibility. This method not only demonstrates good isolation and culture capabilities for the major subtypes of human parainfluenza virus, but also shows good adaptability to clinical samples from different sources and with different initial loads. In addition, this culture system has linear scale-up potential, is not limited by the surface area of ​​adherent culture, and can meet the large-scale virus seed preparation needs from clinical testing to vaccine mass production, providing key technical support for the integrated process from human parainfluenza virus isolation to vaccine production.

The present disclosure provides compositions and methods for manufacturing and obtaining a live attenuated Influenza vaccine (LAIV) composition that can be delivered intranasally to provide protection against influenza virus infection. Said LAIV strains are based on cold adapted, temperature sensitive and attenuated phenotypes of master donor viruses (MDVs) containing the surface glycoprotein genes of the wild type pandemic or seasonal influenza strains. Also, said LAIV strains are further adapted to grow in MDCK cells (Madin Darby canine kidney cells). The use of eggs is avoided in large scale vaccine manufacturing. The purification process is devoid of chromatography steps. The said LAIV composition includes one or more live attenuated influenza vaccine virus and is devoid of polymers and surfactants.

This invention belongs to the field of nanobody technology and provides a method for preparing and applying a single-domain antibody against avian influenza virus H9N2. The application involves using this H9N2 single-domain antibody in the prevention and control of avian influenza virus H9N2. The invention prepares the H9N2 single-domain antibody by immunizing striped bamboo sharks and collecting their tail vein blood. The purity and stability of the H9N2 single-domain antibody were evaluated. The results show that the recombinant single-domain antibody targeting the H9N2 virus HA protein prepared in this invention has high purity and good activity, exhibits good in vitro binding activity with H9N2, and shows good stability at room temperature. The recombinant single-domain antibody 28-H9N2-2SP4H can recognize and bind to the H9N2 virus on the surface of MDCK cell membranes, providing a new direction for the prevention and control of avian influenza as an alternative to traditional antibodies.

This invention relates to the field of bioengineering, and more particularly to sgRNA, MDCK cells, and their applications. This invention provides sgRNA having the nucleotide sequences shown in SEQ ID NO:1 to SEQ ID NO:4. This invention employs a CRISPR-Cas9 gene editing method, which, compared to existing RNAi methods, offers higher specificity and lower off-target risk. Compared to MDCK cells, PCR detection showed a reduction of over 90% in RNA levels in IRF7- / - cells, Western blotting showed a significant decrease in IRF7 protein expression, and viral hemagglutination assays showed higher titers for influenza virus compared to the original cells. Tumorigenicity assays showed that the gene knockout cells retained low tumorigenicity. Compared to existing technologies, the cells of this invention exhibit both low tumorigenicity and significantly increased viral yield, making them suitable for influenza vaccine production.