Provided are a plant EPSPS mutant containing mutations L195P and S247G and an encoding gene and the use thereof, related to the field of genetic engineering technology. Comparing the plant EPSPS mutant with E. coli EPSPS, the amino acid sequence of the plant EPSPS mutant has the mutation L195P at position 195 corresponding to E. coli EPSPS and/or the mutation S247G at position 247 corresponding to E. coli EPSPS. The mutation of either of the two sites or the simultaneous mutation of the two sites can confer or improve the resistance of the plant EPSPS mutant to glyphosate. Plants or recombinant bacteria for transforming the plant EPSPS mutant can grow normally in the presence of glyphosate. The plant EPSPS mutant can not only be used for the cultivation of transgenic crops, but also be used for cultivating non-transgenic plants resistant to glyphosate, such as rice, tobacco, soybean, corn, wheat, cotton and sorghum.
