Abstract: The microsatellite sequences were obtained using a simple sequence repeats (SSR) development method without needing a reference genome and were genotyped through sequencing. The ebwt2InDel software was utilized to identify Insertion/Deletion (InDel)-containing sequences from the whole genome sequencing data of two samples. Using MISA software, 111 perfect SSR-containing sequences were obtained from the InDel-containing sequences by eliminating non-SSR-containing sequences. The SSRgenotyper software was then used to analyze the polymorphism of 30 M.nipponrnse samples, successfully genotyping 19 out of the 111 SSR-containing sequences with a genotyping success rate of 17.12%. Successful genotyping was achieved at 19 SSR loci, all exhibiting 3-base repeats ranging from 4 to 6. Among them, the highest proportion of repetitive sequences was observed with the ATC/ATG unit, accounting for six loci (31.58%). Two of the 19 SSR markers had low polymorphism information content, and 17 had moderate polymorphism information content. These 19 microsatellite markers helped to evaluate the genetic parameters of freshwater shrimp populations and develop freshwater shrimp germplasm resources.