Allele frequency of the G→T mutation of the Col1A1 gene analyzed by an ARMS-PCR in osteoporotic subjects with femoral neck fractures

The allele frequency of the G-->T mutation of COL1A1 gene (collagen type I alpha 1 gene, GenBank accession n. AF017178) was analyzed by a new ARMS-PCR method in 240 osteoporotic subjects bearing a femoral neck fracture. The method is based on the amplification refractory mutation systempolymerase chain reaction (ARMS-PCR). Normal and mutated alleles were detected by two PCRs, in which a common forward primer (positions 1307 to 1336 of the gene) and two reverse primers (positions 1566 to 1546), differing in the 3'-base (3'-C for the normal S allele and 3'-A for the mutated s allele) are used. In the SS condition, amplification occurs only in one of the two PCRs, and in the ss condition only in the other. In the Ss condition both reactions give a product. This ARMS-PCR method avoids the use of any restriction enzyme, as described by Grant and colleagues in a previously published method based on a mismatched reverse primer which introduced a restriction site in the T-substituted (s) allele and in a method recently proposed by Vinkanharju and co-workers. In a survey for COL1A1 polymorphism in 240 osteoporotic subjects with femur fractures, here presented, a frequency of 80.6% was found for the G allele and 19.4% for the T allele. There were 66.7% dominant SS subjects, 27.9% Ss heterozygotes and 5.4% ss recessive homozygotes.