【Objective】To study the rare alleles frequencies and sequences of Expressmarker 22 kit in Guangdong Han Population.【Methods】3495 Samples from unrelated individuals in Guangdong Han Population were screened by using AGCU Expressmarker 22 kit（EX22）and ABI 3100 Genetic Analyzer. Then analyzed the frequencies of the off- ladder（OL）alleles and sequenced the rare alleles obtained based on comparison with the STRBase database and litera? ture.【Results】33 off-ladder alleles with 25 rare alleles were found in 10 STR loci，and allele frequencies ranged from 0.0003~0.0046. Sequencing of the11unreported rare OL alleles showed that most of them have incomplete repeats. 【Conclusion】Off-ladder alleles especially the rare alleles are helpful to improve the power of discrimination and the power of exclusion and to provide samples which will be added its allele into ladders These OL-alleles will supplement forensic DNA database.