Site-directed Mutagenesis Of Southern Rice Black-streaked Dwarf Virus P9-1 And Its Interaction With Small Molecules Of Compounds

Southern rice black-streaked dwarf virus belongs to the genus Fijivirus of the family Reoviridae. SRBSDV is spherical virus, it has 10 RNA genomes, and the S9-1 was predicted to encode the viral matrix protein. Virus matrix protein is the key to the viral replication and proliferation. Dufulin has a control effect for SRBSDV on field. Therefore, to study the interaction of dufulin with SRBSDV P9-1, it can be the basis for the development of new and efficient antiviral drugs.This study is used the crystal structure of RBSDV P9-1 as the template and through the computer to simulate the crystal structure of SRBSDV P9-1, then the computer molecular docking is used to calculate the binding sites of dufulin with SRBSDV P9-1. The mutant plasmids are obtained by the PCR site directed mutagenesis method, the wild type and mutant of SRBSDV P9-1 are aquired from prokaryotic expression system. Finally, through fluorescence spectra, isothermal titration calorimetry and microscale thermophoresis to study the binding force of dufulin with proteins. Experimental results show that using site directed mutagenesis PCR method can obtain mutant plasmids. The results of the three methods can be seen, the binding force of dufulin with the 175 Arg P9-1 mutation is weaker than dufulin with the wild type. The results of the experiment are in agreement with the results of computer simulation, indicating that SRBSDV P9-1 175 arginine is the key of dufulin binding sites.