MDBK IRF3/IRF7 KNOCK OUT MUTANT CELL AND ITS USE FOR VACCINE PRODUCTION
The present invention pertains to a Madin-Darby bovine kidney (MDBK) cell, wherein the interferon regulatory factors (IRF) IRF3 and/or IRF7 encoding genes are functionally inactivated. The invention also pertains to a cell culture comprising the MDBK cell, use of the MDBK cell culture, a method for the production of a virus using the cell and a vaccine prepared by using the cell.
1 . A Madin-Darby bovine kidney (MDBK) cell, characterized in that the cell is genetically engineered by which the interferon regulatory factors (IRF) IRF3 and/or IRF7 encoding genes are functionally inactivated.
2 . The MDBK cell of claim 1 , IRF3 and IRF7 encoding genes are functionally inactivated.
3 . The MDBK cell of claim 1 , which is infected with a virus.
4 . The MDBK cell of claim 3 , wherein the virus is an enveloped virus.
5 . The MDBK cell of claim 3 , wherein the virus is selected from the group consisting of BRSV, bovine diarrhea virus, bovine rhinotracheitis virus, bovine parvovirus, bovine adenovirus, bovine corona virus and bovine parainfluenza virus.
6 . The MDBK cell of claim 3 , wherein the virus is a bovine respiratory syncytial virus (BRSV).
7 . A cell culture comprising the MDBK cell of claim 1 .
8 . (canceled)
9 . A method for the production of a virus, the method characterized in that it comprises the steps of:
(a) culturing the MDBK cell according to claim 1 ,
(b) infecting the cell with a virus,
(c) allowing the virus to replicate, and
(d) isolating the virus from the cell culture.
10 . A vaccine composition characterized in that it comprises the virus infected cell culture of claim 7 or a virus isolated therefrom and a pharmaceutically acceptable carrier.
11 . A method for the preparation of a vaccine composition comprising a virus, the method characterized in that it comprises the steps of:
(a) culturing the MDBK cell according to claim 1 ,
(b) infecting the cell with a virus,
(c) allowing the virus to replicate,
(d) isolating the virus from the cell culture, and
(e) mixing the virus with a pharmaceutically acceptable carrier.
12 . The vaccine composition of claim 10 , characterized in that the pharmaceutically acceptable carrier comprises a natural deep-eutectic solvent (NADES) having a water activity of less than about 0.8.
13 . The vaccine composition of claim 12 , wherein the virus is a live virus or live attenuated virus.
14 . The vaccine composition of claim 13 , wherein the virus is a bovine respiratory syncytial virus (BRSV).
15 . A method of CRISPR-Cas9 mediated gene editing, characterized in that the method comprises the steps of:
(a) providing an MDBK cell and
(b) performing CRISPR-Cas9 mediated functional inactivation of the interferon regulatory factors (IRF) IRF3 and/or IRF7 encoding genes of the MDBK cell.
16 . A method for the production of a virus, the method characterized in that it comprises the steps of:
(a) culturing the MDBK cell according to claim 1 ,
(b) infecting the cell with a virus, and
(c) allowing the virus to replicate.
17 . A method for the preparation of a vaccine composition comprising a virus, the method characterized in that it comprises the steps of:
(a) culturing the MDBK cell according to claim 1 ,
(b) infecting the cell with a virus,
(c) allowing the virus to replicate, and
(d) mixing the cell culture infected with the virus with a pharmaceutically acceptable carrier.
18 . The method of claim 17 , characterized in that the pharmaceutically acceptable carrier comprises a natural deep-eutectic solvent (NADES) having a water activity of less than about 0.8.