Structural And Functional Analysis Of Baculovirus P10 Gene

In the late phase of baculovirus infection, there are large bundles of fibrillar structures existing in the cytoplasm and nucleus of infected cells. Immuno-gold electron microscopy had revealed that these fibrillar structures were composed predominantly of viral encoded protein with molecular mass of approximately 10kDa. Therefore, the gene encoding this protein was named as p10. Previous studies suggested that p10 is not essential for virus replication, but it might play roles in stabilizing polyhedra, releasing polyhedra and nuclear membrane disintegration. Now p10 is found in 21 of 29 sequenced baculovirus, suggesting the conservation of p10 in baculovirus evolution. In this thesis, the structure and function of p10 gene of HaSNPV and AcMNPV are studied.Chapter 1 is a general introduction. It contained a introduction of biology, molecular biology and application of baculovirus. The state of the art of baculovirus p10 research was introduced and the aim of the thesis was presented.In chapter 2, the HaSNPV p10 gene was cloned and expressed in bacteria cells with GST-tag. The GST-P10 protein was purified by sepharose 4B affinity beads and immunized into rabbits to generate antiserum against HaSNPV P10. Western blot and immuno-gold microscopy revealed that the obtained antiserum reacted with HaSNPV P10 specifically.In chapter 3, the functional domains required for HaSNPV P10 filament formation were identified. We have tried to identify the functional domains for the P10 filament formation by green fluorescence protein (GFP) tag. Different lengths of HaSNPV p10 gene were constructed with GFP fused in frame to the C-terminus. The constructs were transfected into insect and mammalian cells and observed by confocal microscopy. The results indicated that the first N-terminal 66 amino acids, which form the complete...