الفهرس 
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Abstract
Norovirus is the leading cause of nonbacterial gastroenteritis all over the world .In developing countries, it accounts for 200000 deaths in children younger than 5 years annually. They belong to family Caliciviridae and contains at least six genogroups with more than 30 genotypes. But only strains from genogroups I, II and, IV infect human, where NoV GII predominate. Noroviruses have single stranded RNA positive sense genome of approximately 7.5 kb in three ORFs. ORF2: encodes the major viral capsid protein and the antigenic determinant (VP1). VP1 is composed of shell (S) and protruding (P) domains. Shell domain is known for relative genetic and antigenic conservancy.
The wide genetic and antigenic variation even among the same genogroup renders the species, generally, elusive for sensitive specific diagnosis and vaccine design. Virus uncultivability and lack of a suitable animal model contributed to failure of development of standard sensitive economic diagnosis and commercial vaccine. To overcome uncultivability, several trials for norovirus heterologous protein expression was performed. But all of them showed drawbacks that relates to design of the expressed protein, or to limitations of the utilized host.
In this study we designed a multifunctional chimeric construct. It encodes Shell (S) domain of HuNo G.II.4 and NDV fusion (F) domain. Both domains were linked using proper peptide linker. Finally a stabilizer was linked to the C-terminus of the whole construct. The stabilizer was added to enhance the expression of construct in the newly emerging algal system; Chlamydomonas reinharditti But fusion domain was added to ease purification of produced protein by affinity.
Selected shell (S) was verified for antigenic conservancy among large group of G.II.4 strains and their contents of T- Cells epitopes. We tested the effect of both linker and stabilizer on the 3D structure of both domains. To, in silico, verify safety of the protein, we confirmed that protein sequence is free from allergic and toxic motifs. Conservation of glycosylation sites and subcellular localization were also validated.
The verified construct was used to produce a transgenic C. reinharditti strain- it was selected as host because of easiness of cultivation and manipulation; rapidity of growth; safety; and economics of production. For detection of transgenic colonies, we devised a new, economic, simple, and rapid method for extraction of the genomic DNA of C. reinharditti different strains.
The produced transgenic C. reinharditti strain is supposed to be used for robust production of GⅡ.4 shell domain that can be easily purified by affinity. Besides C. reinharditti aforementioned merits, robust expression and easy purification are supposed to render the expressed protein amenable for economic, safe, broader, efficient vaccine development. The recombinant pure protein can pave the way for economic improvement of kits for indirect (virus specific antibody) diagnosis for epidemiological investigation. Moreover, the newly devised genomic extraction method is an asset to algae biotechnology labs in basic and applied research.