A RAPID METHOD TO STUDY HEAT-SHOCK PROTEIN-70-2 GENE POLYMORPHISM IN INSULIN-DEPENDENT DIABETES-MELLITUS

To examine the role of DNA loci within the human leukocyte antigen (HL A) region and insulin-dependent diabetes mellitus (IDDM), we studied f ine mapping of HSP70-2 gene. Polymerase chain reaction (PCR)-based gen otyping was then developed and applied to type HSP70-2 in 59 patients with IDDM and 83 unrelated controls recruited from the inhabitants of northern Taiwan. Southern blot analysis revealed a diallelic PstI poly morphism of the HSP 70-2 gene, i,e., 9.6- and 8.5-kb alleles, The poly morphic site was mapped in the intragenic PstI sequences (nucleotides 1051-1056) of the HSP70-2 gene. PCR-based restriction fragment length polymorphism studies revealed that the frequency of the 8.5-kb allele was increased in IDDM (56.8%, vs, 40.4% in controls; p < 0.009), with a relative risk of 1.93 (95% confidence interval = 1.20-3.11). The gen otypic frequencies of 9.6/9.6, 9.6/8.5, and 8.5/8.5 were 17.0, 52.5, a nd 30.5% for IDDM were different from these of controls (36.1, 47.0, a nd 16.9%, respectively; the homozygous 9.6/9.6 genotype was significan tly decreased in the IDDM group, p < 0.02). In conclusion, we provide a simple, rapid, and nonradioactive method for HSP70-2 genotyping. Our data confirmed that the 8.5-kb allele of HSP70-2 was associated with IDDM susceptibility in the Taiwanese population.