Mutations in type I collagen genes in Japanese osteogenesis imperfecta patients.

Background: Osteogenesis imperfecta (OI) is an autosomal dominant disorder of connective tissue characterized by bone fragility and low bone mass. COL1A1 and COL1A2 genes are very large and have been rarely analyzed systematically in Japan. The aim of this project was to develop an effective and convenient method of finding mutations in the COL1A1 and COL1A2 gene by using denaturing high-performance liquid chromatography (DHPLC).

Methods: Polymerase chain reaction (PCR) amplicons of genomic DNA from the COL1A1 or COL1A2 gene were followed by heteroduplex analysis by DHPLC. Products containing heteroduplexes were then sequenced.

Results: Twenty-two OI families were analyzed, and 193 of the 1122 PCR products in the COL1A1 gene, all containing heteroduplexes, were sequenced. Sixty-two samples had single-base substitutions or single-base deletions or insertions within introns. Eight had single-base substitutions in exons. Six were pathogenic mutations, and two were silent mutations. In 16 families not identified with pathogenic mutation in COL1A1, COL1A2 was similarly analyzed. A total of 138 of the 848 PCR products were sequenced, and 46 samples had single-base substitutions, or single-base deletions or insertions within introns. Twenty-four samples had single-base substitutions in exons. Three were pathogenic mutations and the others silent.

Conclusions: Mutations were identified in nine COL1A1/COL1A2 associated with OI type I-IV genes by scanning with DHPLC. Software was used to detect point mutation and large deletions/insertions in COL1A1 and COL1A2 genes.