Comparison Of Buzura Suppressatia Nucleopolydrovirus (busunpv)isolates And Its Detection Method

The thesis made the test by biological in order to know the virulence of different strains of Buzura suppressatia nucleopolydrovirus, to survey external characteristics of different strains with scanning electron microscopy and distribution of virus particles with transmission electron microscope by production of thin sections; virus genome restriction enzyme analysis of viral nucleic acid changes. To design specific primers according to the polyhedrin envelope protein gene (pep) and lef-2 gene of BusuNPV, the PCR detection technology of BusuNPV successfully established. The study and results are as follows:(1)There is no significant difference virulence of all BusuNPV isolates by measured. The LC50 of LF, SF, and WF were 4.39×104, 6.65×104 and 1.02×105PIB/ml; the LT50 were 5.59, 5.69, and 5.17 d. It is showed that all isolates are single-particle-embedded nuclear polyhedrosis virus by observing photographs of scanning electron microscopy and transmission electron microscope, polyhedron diameter is 0.85-1.50um, the virus particle size is 0.195-0.246 um×0.054-0.072 um.(2)Digesting the whole genome sequence of BusuNPV by selecting 5 kinds of enzymes. In addition to EcoRⅠ,endonuclease digestion of foreign bands are small, molecular weight, which is difficult to comprising the isolate. Digestion of DNA fragments by EcoRⅠwas about 12, the molecular weight was less than 15kb, there is no differences in isolates. Digesting the BusuNPV by double enzyme genome sequence, DNA was fully digested bands are more suitable for comparison in isolates. However, restriction enzyme digestion patterns observed, the isolated enzyme band number and size have no different.(3)Identifying two specific gene fragments pep and lef-2 of BusuNPV by comparing gene, and designing two pairs of specific primers BPF and BLA. Establishing a rapid detection BusuNPV method based PCR technology.(4)PCR detection technology of BusuNPV is not only detecting BusuNPV from the eggs and the pupae successfully within a short time ,but also its sensitivity was 1 fg/mlDNA. After the template diversity, polyhedral suspension can also be used directly as template for PCR technology to detect BusuNPV. (5)Established PCR technology in experiment was rapid, specific, sensitive, simple and the application results was very good. It can be used to study the proliferation disciplinarian of the virus in the insect body and the detecting situation of virus in the insect body in forest.