[Screening mutations in phenylketonuria by means of nonradioactive reverse dot blot hybridization].

OBJECTIVE

To establish a simple, accurate and rapid method for screening of the mutant genes in phenylketonuria (PKU).

METHODS

Four exons harboring the mutations, Y204C (exon6, E6), R243Q(E7), Y356X(E11) and R413P(E12), were amplified by polymerase chain reaction (PCR) with incorporation of biotinylated deoxynucleotide(biotinylated-11-dUTP or biotinylated-14-dCTP). Hybridization between immobilized allele-specific oligonucleotide probes and biotin-labelled amplified DNA was performed and nonradioactively detected by a colorimetric reaction using streptavidin-alkaline phosphatase.

RESULTS

The methods of non-radioactive reverse dot blot hybridization were established to screen the mutations. We detected the genotypes of five PKU patients and found that three of them carried R243Q mutation and one of the three also carried Y356X mutation. These results were confirmed by PCR-single strand conformation polymorphism.

CONCLUSIONS

This method is suitable for rapid screening for common mutations in Chinese PKU patients.