OBJECTIVE To figure out the polymorphism of three Y-STR loci in isolated populations and explore the consanguinity of the populations with the use of Y-STR. METHODS Male samples were selected from two isolated populations(80 and 60 males) in Zhejiang province and one open population (36 males), genescan was performed with males' DNA by genescan technology with ABI PRISM 377 sequencer at Y chromosome loci DYS388, DYS390 and DYS395. RESULTS DYS388, DYS390, DYS395 allele counts in Yushan island population, Taohua island population and open population were 8, 9, 7, 5, 6, 7 and 6, 6, 5 respectively. Gene diversity was between 0.70-0.80 in the three populations. There was no difference in distribution of allele frequency and shared genotypes between the isolated populations and the open population by statistical test. Genetic distance is long between Taohua island population and open population, short between Yushan island population and open population, and moderate between Yushan island population and Taohua island population. CONCLUSIONS The main allele is 129 at DYS388; 215 at DYS390; and 119 at DYS395. The distribution of allele frequency and gene diversity at DYS388, DYS390, DYS395 loci, and the shared genotypes between populations as well as the genetic distance are unable to explain the blood relationship between the isolated and open populations, suggesting the additional studies in large sample size will be necessary to use Y-STR for exploring the blood relationship between populations.