Abstract: The rabies virus strain GX074 was isolated from the brain of a healthy dog in the Guangxi Province in 2003, and the amino acids at positions 253, 263, and 273 of its G protein were replaced to the corresponding sites of the attenuated vaccine strain rRC-HL by reverse genetic technology to obtain the recombinant strain Mu11. To verify the effects of amino acids at positions 253, 263 and 273 of G protein on the growth characteristics of rabies virus, this study used the infectious cDNA cloning plasmid of Mu11 strain as the basis, and restored the amino acid mutations at positions 253, 263, and 273 of strain Mu11 G protein to the corresponding amino acids sites of the parental attenuated strain rRC-HL, and the mutant virus strain Mu11^R253.263.273 was obtained. It was confirmed that the mutant sites of the strain Mu11^R253.263.273 were correct by indirect immunofluorescence assay and gene sequencing experiment. By measuring its viral titer and multi-step growth curve, it was demonstrated that the mutant virus strain Mu11^R253.263.273 had stable genomeand its replication ability was consistent with the parental attenuated strain rRC-HL in cells. These studies indicate that amino acids 253, 263, and 273 of rabies virus G protein have an important effect on the growth characteristics of the strain Mu11 and the parental virulent strain GX074, which provides scientific basis for further research on its pathogenicity.