An autosomal recessive genetic disorder characterized by phenylalanine hydroxylase(PAH) deficiency, classic phenylketonuria (PKU) may lead to irreversible mental retardation, if left untreated. The prevalence of PKU in China is about 1 in 16,500. In our study, we analyzed the polymorphism of tetranucleotide Short Tandem Repeat(STR) sequence in intron 3 of the PAH gene in 52 normal individuals and 46 members of 23 PKU families. Nine alleles were identified, with a continuous distribution from 224 bp to 256 bp. The smallest(224 bp) was detected in Chinese population for the first time. The polymorphism information content was found to be 0.730 in PKU families as distinguished from 0.654 in normal population. This single marker is nearly twice as informative as RFLPs haplotype analysis in China as reported by others. Next, using a simple and rapid method we reported formerly (Fang Song et al. Chinese Science Bulletin. 39(1), 1994) -- Multiplex Allele-specific PCR (MASPCR), five known mutations (R111X, Y204C, R243Q, Y356X, R413P) were detected in the same 23 PKU families which accounted for about 47% of all PKU alleles, with the most frequent mutations being R243Q and R413P. Capitalizing on these findings, we devised a sensitive combined approach to diagnose PKU, which has proved to be a simple, effective and accurate. The families which can be diagnosed by lab approach have risen from 52.2%(STR) and 26.1%(MASPCR) respectively to 78.3%(Combined Approach). And the cases that were diagnosed by exclusion were reduced to 21.7% as distinguished from 43.5%(STR) and 43.5%(MASPCR). As a result, the families that can not be diagnosed at all were 0% as against 4.3%(STR) and 30.4%(MASPCR). The prenatal diagnosis of 6 fetuses at risk were detected with this combined approach and were confirmed after birth or by examination of aborted materials.