A new method of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was developed for genetic typing of a point mutation of the Bruton's tyrosine kinase (Btk) gene in CBA/N mice bearing an X-linked recessive immunodeficiency (xid). Since restriction site useful for RFLP analysis does not exist in the spontaneous mutant Btk^xid locus, an artificial restriction site was introduced by PCR amplification with a modified primer. The five genotypes of the Btk locus (Btk^xid/ Btk^xid, Btk^xid /Btk⁺ and Btk⁺/Btk⁺ females and Btk^xid/Btk^null and Btk⁺/Btk^null males) could be distinguished by three patterns clearly and easily. This PCR-RFLP analytic method will be useful as a tool in the production of congenic mice and mice with multiple immunodeficient genes.