Prokaryotic Expression Of Rotavirus Structure Protein And In-Vitro Assembly Of Virus-like Particles

Rotavirus is the most common cause of severe gastroenteritis among children worldwide,Almost every child has Infected rotavirus once or more before 5 years old, resulting in approximately 200 million diarrhea cases,more than 2 000 000 hospitalization and 1 million death every year worldwide,the healthcare cost of rotavirus is over 1 billion dollars every year in the United States.A lot of research on rotavirus has been under way since it was discovered in human in 1973,we understand more deeply in the structure and infection mechanism of the virus,but there is no specific medicine to rotavirus diarrhea,thus quick, accurate diagnostics and safe,effective vaccines are important in rotavirus prevention and control.VLPs has great Opportunity as a rotavirus candidate vaccine and is important in the research of both infection and assembly mechanism.First,we expressed the structure protein VP2 in E.coli and assembled it into CLPs in-vitro for the first time,we also studied the role of buffers and additives on its stability,and it laid the foundation for the study of VP2 immunogenicity and protectivity;Second,We expressed the inner layer protein VP6 and its deletion mutants,we found that 243-334aa is important for VP6 trimerization,at the same time, we expressed single point mutants of VP6 and found Cys197 is important for VP6 polymerization,the WB results show the influence of deletion and point mutation on VP6 immuno-reactivity;third,we constructed VP4 and VP7 expression plasmids; furthermore,we prepared mAb to rotavirus,mainly to VP6 and primary results show that 7H11,2A9 and 5C2 are excellent in both sensitivity and specificity.This study laid the foundation for rotavirus VLP in-vitro assembly and vaccine development,it is important for the study of rotavirus infection and assembly mechanism.