|Budget Amount *help
¥2,000,000 (Direct Cost : ¥2,000,000)
Fiscal Year 1992 : ¥600,000 (Direct Cost : ¥600,000)
Fiscal Year 1991 : ¥1,400,000 (Direct Cost : ¥1,400,000)
We have investigated the genetic polymorphisms of immunoglobulin G (IgG) and immunoglobulin kappa light chain (Ig kappa) genes by using the polymerase chain reaction (PCR).
In the present methods, samples of extracted human genome DNA were subjected to the first PCR for the amplification of the polymorphic regions within the immunoglobulin genes.
Small portions of the amplification products obtained from the first PCR were further subjected to the second PCR amplifications in which the allele specific primer sets were used for genotyping of the immunoglobulin genes.
By this methods, alleles on the IgG3 constant gene, G3m*t andnon-G3m*t alleles, which determine the G3m(t) allotype and its antithetical isoallotype non-G3m(t), respectively, and alleles on the Ig kappa constant gene, Km*1, Km*1.2 and Km*3, were examined for genotyping.
For the examination of the Ig kappa constant gene, 353bp fragments which contain the polymorphic region of the Km allotypes were successfully amplified from sample DNAs by the first PCR. The second PCRs using the products of first PCR as samples could clearly identify the sequence changes between the three alleles. Results of a tiny population study among 72 individuals living in Okayama Prefecture showed the gene frequencies of Km*3= 0.736 and Km*1.2= 0.264.
In the case of the examination of the IgG3 constant gene, 726bp fragments which contain the polymorphic region of the G3m(t) allotype were successfully amplified from sample DNAs by the first PCR. Using the products of first PCR as samples, the second PCRs could generally distinguish the nucleotide substision between the two alleles.