Objective To use random amplified polymorphic DNA (RAPD) for species identification and origin determination of imported rodents. Methods Genetic DNA of rodents was extracted from tail tissue specimens and amplified by RAPD. Genetic polymorphisms in rodents were distinguished by differences in strong RAPD banding patterns. Results A quick DNA preparation method for sewer rats and their genetic maps were established. In accordance with different polymorphisms of 120 primers, a set of universal primers for sewer rats were synthesized, designated P1 (5'-TCG GCG GTT C-3'), P2 (5'-GAG AGC CGT C-3'), P3 (5'-ATG GCA TCT C-3'), and P4 (5'-CTT GGC ACG A-3'). Amplification of a target fragment (925 bp) in sewer rats using the universal primers all yielded good results. Conclusion Analysis of the degree of homology between individual DNA sequences and determination of their genetic distance allow identifying the origin of rodents as imported or not. The obtained data provide a reference for tracing the dispersal path and history of rodents and detecting early-warning signs of rodent-borne diseases.