Abstract: Microcystis is the most common cyanobacteria in China’s freshwater, which has a great impact on ecological environment and urban domestic water. In order to explore the genetic diversity and phylogenetic relationship of Microcystis in Yuqiao Reservoir, MLST (Multilocus sequence typing) gene pool of the Microcystis of Yuqiao Reservoir was constructed based on 7 housekeeper genes (ftsZ, glnA, gltX, gyrB, pgi, recA and tpi) of 10 strains, meanwhile, the synthesis gene mcyE, cyanobacteria characteristic gene, cyanobacteria blue protein cpcBA gene and 16S rDNA gene of microcystins were determined. Based on the Maximum likelihood (ML) phylogenetic tree, we compared STs of this study with STs in Poyang Lake and Japanese Lake, and found STs of Yuqiao Reservoir formed an independent branch. Phylogenetic analysis indicated that ten unique sequence types (STs) had high genetic diversity with a H value of 0.938 in Microcystis. Phylogenetic analysis showed that the existing Microcystis in Yuqiao Reservoir had the same ancestor and relative origin. The formation of Microcystis group structure in Yuqiao Reservoir may be caused by the combination of genetic drift of environmental selection and homologous recombination.