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CN-121971612-A - Application of LRP1 protein or inhibitor of coding gene thereof in preparation of MDV infection resisting preparation

CN121971612ACN 121971612 ACN121971612 ACN 121971612ACN-121971612-A

Abstract

The invention relates to the technical field of genetic engineering, and discloses application of an inhibitor of LRP1 protein or a coding gene thereof in preparation of an MDV infection resistant preparation. In particular to an application of LRP1 protein or a coding gene thereof in regulating and controlling Marek's Disease Virus (MDV) in vitro infected Chicken Embryo Fibroblasts (CEF). The invention discovers that knocking down the LRP1 gene in CEF cells can extremely obviously reduce MDV infection rate, and further provides application of the LRP1 protein or the coding gene thereof or biological materials containing the coding gene thereof in resisting MDV infection. The invention defines the function of LRP1 gene in MDV in vitro infection CEF, and lays a foundation for further researching the antiviral mechanism of LRP1 gene and the application of LRP1 gene in disease-resistant breeding.

Inventors

  • LIAN LING
  • ZHENG GANG
  • YANG YUQIN
  • Du Yushuang
  • WEI YUNJIE

Assignees

  • 中国农业大学

Dates

Publication Date
20260505
Application Date
20260106

Claims (9)

  1. Use of an inhibitor of lrp1 protein or a gene encoding the same in the preparation of a formulation against MDV infection; Wherein, the reference sequence number of the mRNA sequence of the LRP1 coding gene from the chicken in NCBI is NM_205242.3.
  2. 2. The use according to claim 1, wherein the inhibitor is a substance capable of inhibiting the expression or activity of LRP1 encoding gene or LRP1 protein at the transcriptional or translational level; Preferably, the inhibitor is selected from at least one of shRNA, siRNA, dsRNA, miRNA, cDNA, antisense RNA/DNA, low molecular compounds, peptides, antibodies.
  3. 3. The use according to claim 2, wherein the inhibitor is a nucleic acid molecule targeting the LRP1 encoding gene, the function of which is to reduce the expression level of the LRP1 encoding gene in a cell.
  4. 4. The use according to claim 2, wherein the inhibitor is an siRNA comprising a sequence selected from any one of or a combination of LRP1-si-A, LRP 1-si-B: LRP1-si-A: Sense strand CGAGAUGGUUUACCUCAAUTT; the antisense strand AUUGAGGUAAACCAUCUCGTT; LRP1-si-B: sense strand GCCGCUGCAUAAACAUCAATT; Antisense strand UUGAUGUUUAUGCAGCGGCTT.
  5. 5. The use according to claim 4, wherein the siRNA is expressed by a recombinant expression vector comprising a promoter capable of driving expression of the siRNA in a host cell and a coding sequence for the siRNA.
  6. 6. The use according to claim 5, wherein the backbone vector of the recombinant expression vector is pGPU6/Neo.
  7. 7. An application of screening target points of anti-MDV drugs, which is characterized in that the target points are LRP1 proteins or encoding genes thereof.
  8. 8. A method for creating MDV-resistant transgenic chicken or gene-editing chicken is characterized in that the genome of the chicken is modified by adopting a genetic engineering technology, so that the functional expression of LRP1 genes in the chicken is reduced.
  9. 9. The method of claim 8, wherein the method of genomic modification is selected from the group consisting of gene knockout, gene knockdown, and introduction of a mutation that results in loss of function in the chicken endogenous LRP1 gene.

Description

Application of LRP1 protein or inhibitor of coding gene thereof in preparation of MDV infection resisting preparation Technical Field The invention relates to the technical field of genetic engineering, in particular to application of an inhibitor of LRP1 protein or a coding gene thereof in preparation of an MDV infection resistant preparation. Background Chicken Marek's Disease (MD) is a lymphoproliferative Disease caused by Marek's Disease Viurs, MDV infection, and induces acute or subacute lymphocytoma after virus infection, causing serious immunosuppression, which is one of the important prevention and control diseases threatening the healthy cultivation of poultry industry. MDV is a strictly cell-binding virus that is hardly free from extracellular presence, except for intact virions that are visible in the feather marrow of infected chickens. In the in vitro breeding process of conventional viruses and vaccine viruses, primary chick embryo fibroblasts (Chicken Embryo Fibroblasts, CEF) are often used as virus vectors, and have the advantages of easy acquisition, strong proliferation capacity and high infection rate, and are widely applied to vaccine development and virology related research, so CEF is also a good cell model for researching virus infection. Low density lipoprotein receptor-related Protein-1 (Low density lipoprotein Receptor-related Protein 1, lrp 1) is a multifunctional transmembrane receptor belonging to the LDLR (Low density lipoprotein Receptor) family, which is involved in a variety of cellular processes including endocytosis, intracellular signaling, lipid homeostasis, and apoptotic cell clearance, among others. LRP1 is also reported to be an invading factor of the Rift Valley Fever Virus (RVFV) and a key receptor for invasion of host cells by the febrile associated thrombocytopenia syndrome virus (SFTSV), whose own functions include participation in endocytosis, endocytic receptors as phagocytosis of apoptotic cells, participation in lipid homeostasis, participation in intracellular signaling, and the like. Disclosure of Invention The object of the present invention is to provide the use of inhibitors of LRP1 protein or genes encoding the same for the preparation of a formulation against MDV infection. In the invention, the LRP1 gene is identified to participate in the MDV infected cell process, the effect of the LRP1 gene in MDV in-vitro infection is clear, and targets can be provided for leading-edge researches of developing Marek's disease resistance gene editing chickens and the like. To achieve the object of the present invention, in a first aspect, the present invention provides the use of an inhibitor of LRP1 protein or a gene encoding the same for the preparation of a formulation against MDV infection. Wherein, the reference sequence number of the mRNA sequence of the LRP1 coding gene from the chicken in NCBI is NM_205242.3. Further, the inhibitor is a substance capable of inhibiting the expression or activity of an LRP1 encoding gene or LRP1 protein at the transcriptional or translational level; Preferably, the inhibitor is selected from at least one of shRNA, siRNA, dsRNA, miRNA, cDNA, antisense RNA/DNA, low molecular compounds, peptides, antibodies, etc. Further, the inhibitor is a nucleic acid molecule targeting the LRP1 encoding gene, which functions to reduce the expression level of the LRP1 encoding gene in a cell. Preferably, the inhibitor is an siRNA comprising a sequence selected from any one of or a combination of LRP1-si-A, LRP 1-si-B: LRP1-si-A: Sense strand CGAGAUGGUUUACCUCAAUTT; the antisense strand AUUGAGGUAAACCAUCUCGTT; LRP1-si-B: sense strand GCCGCUGCAUAAACAUCAATT; Antisense strand UUGAUGUUUAUGCAGCGGCTT. Further, the siRNA is expressed by a recombinant expression vector comprising a promoter capable of driving expression of the siRNA in a host cell and a coding sequence for the siRNA. Preferably, the starting vector of the recombinant expression vector is pGPU6/Neo. In a second aspect, the invention provides an application of screening a target of an anti-MDV drug, wherein the target is LRP1 protein or a coding gene thereof. In a third aspect, the present invention provides a method of creating an anti-MDV transgenic chicken or a genetically engineered chicken, by modifying the genome of the chicken using genetic engineering techniques, so as to reduce the functional expression of the LRP1 gene in the chicken. Further, the method of genomic modification may be selected from any one of gene knockout, gene knockdown or introduction of mutation causing loss of function of the chicken endogenous LRP1 gene. In a fourth aspect, the invention provides the use of LRP1 protein or a gene encoding the same for modulating MDV infection in vitro. By means of the technical scheme, the invention has at least the following advantages and beneficial effects: The invention constructs two siRNAs capable of effectively knocking down chicken LRP1 expression and corresponding