Designing primer for exon 2 of HLADRB1 gene in type 1 diabetes / Noor Syafiqah Rosli

Type 1 diabetes is known as a complex disease as several gene mutations are likely to contribute to the disease. The HLA region on chromosome 6p21, which includes the major histocompatibility complex (MHC) class II has been identified as the major genetic determinant for the development of Type 1 diabetes. It has been identified that MHC class II genes DRBl, DQAl, and DQBl are the loci of greatest genetic risk for this disease. The strongest association of these genes with Type 1 diabetes were observed with DRBl and DQBl genes. DRBl gene was found to influence the risk ofDQBl allele susceptibility. Therefore, HLADRBl was chosen in this research. The objective of this research is to design a set of primers to amplify exon 2 of HLADRB 1 gene using Polymerase Chain Reaction (PCR) technique. A set of primer was designed for the amplification of exon 2 of HLADRB 1 gene (NG_029921) by using NCBI Primer BLAST The primers together with the PCR components were assembled for PCR amplification and optimization which then subjected to gel electrophoresis. The desired amplicons were sent for DNA sequencing and analysis of SNPs were conducted. In the results, seventeen SNP were found for this exon; 9 synonymous and 8 non-synonymous. Deletion of GC at position rs13383 and rs13384 result in the conversion of Serine to Lysine. Insertion of Cat 350nt causes a conversion of amino acid from Serine to Threonine whereas at 36lnt, insertion of C causes conversion of Glycine into Arginine. In conclusion, the newly designed primer set was able to amplify exon 2 of HLADRB 1 gene successfully. However, all SNPs were not reported as clinically significant in the previous reports.