الفهرس 
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Abstract
In the current study, construction of platform for delivery of mRNA expressing fusion (F) protein of NDV was conducted. The construction of recombinant mRNA containing the fusion gene of NDV followed by expression and characterization of the fusion protein in vitro and in vivo was achieved by the following steps:
1. Construction of mRNA-based constructs: by Cloning of fusion (F) gene of LaSota (genotype II) and NDV genotype VII strain into the pVAX1 vector.
2. Production of mRNA vaccines by; In vitro transcription, capping of mRNAs, Complexing of mRNA and CP 2k and characterization of CP 2k/mRNA by agarose gel electrophoresis
3. In-vitro characterization of CP 2k/mRNA vaccines by; Immunofluorescence assay (IFA) and Western Immunoblotting assay.
4. In vivo immunogenicity of CP2k/mRNA NDV vaccines through Animal experiment and inoculation of mRNAs constructs into mice, Evaluation of cytokines responses by RT-qPCR; Determination of antibody immune response by HI assay and Histopathology and immunohistochemistry.
Vaccination of domestic poultry may help to control and eradicate viral infection and to reduce economical losses in poultry sectors. Availability of high performance and protective vaccine is an urgent demand to attain control of most important respiratory viruses especially Newcastle disease. To generate a novel approach in the development of new vaccine, the vaccine should induce combined humoral and cellular immunity resulting in broad protection against antigenically drifted strains. mRNA based vaccines represent an innovative immunization modality overcoming the drawbacks of conventional and subunit vaccines. mRNA vaccines are highly versatile, non-toxic molecules that are easy to produce and store, its ability to promote broader cytotoxic T-cell responses and induce efficient humoral and cellular immunity resulting in wider and longer protection. Therefore; the proposed study is directed to construct nanopolymer (CP2K) based mRNA vaccine against Newcastle disease virus that can act as both vaccine delivery vehicle and adjuvants and will enhance both humoral and cellular immunity with extend the half-life of antigens ensuring a higher antigen exposure to APCs after vaccination. CP2K is a cationic polymer consisting of β-cyclodextrin and low-molecular-weight PEI 2k, which carries the high mucosal affinity of cyclodextrin and the good adjuvanticity of the cationic PEI polymer. Our study provides in vivo evidence that CP polymer administered intranasally efficiently delivered an mRNA vaccine across the nasal epithelial barrier and induced potent immune responses against Newcastle disease virus F protein without causing absorption of toxins present in the nasal cavity. Intranasal inoculation of mice with CP 2k/mRNA led to strong mucosal and systemic immune responses in a balanced Th1/Th2 profile. Using CP2k, condensation of mRNA into particles also increases the expression of the antigen encoded in the mRNA and therefore higher antigen-specific immunity against F protein. Thus, this cationic CP polymer represents an excellent platform material for intranasal mRNA vaccination.