Analysis of the mutation of exon 8 and 12 of Wilson's disease gene

Objective To sequence and study the exon 8 and 12 of hepatolenticular degeneration(Wilson's disease,WD) gene and to establish the way in direct gene diagnosis. Methods To extract the genomic DNA of 102 WD patients and 82 normal controls,and amplify exon 8 and 12 of ATP7B gene by polymerase chain reaction(PCR). To analyze the amplification of exon 8 and exon 12 by digestion with MspI and Tail respectively before sequencing amplification of all the patients and normal controls. Results Digested by MspI through amelioration,35 cases were abnormal. Sequence results showed that 34.31%of the cases had homozygous or heterozygous Arg778Leu mutation in exon 8,among which 12 were found to have Leu770Leu polymorphism simultaneously;13 cases were abnormal using digestion by Tail,subsequent direct sequencing showed 12.75% of the cases had heterozygous Thr935Met mutation in exon 12,one case was identified as heterozygous Arg919Gly,and Arg952Lys was detected in patients and normal control. Conclusion Arg778Leu in exon 8 and Thr935Met in exon 12 are the hot point mutation of ATP7B gene,and PCR-MspI and PCR-Tail digestion are reliable for molecular diagnosis in patients with Wilson's disease.