The PTPN22 gene plays vital role in T1D onset by encoding Lymphoid-specific phosphatase (LYP) that lead to T-cell receptor-associated CsK kinase inactivation and preventing T-cell spontaneous activation by dephosphorylation. VDR gene encoded for VDR receptor is involved indirectly in prevention of T1D onset by promoting insulin production. The present study was conducted to determine the mutations on rs2476601 and rs1544410 polymorphic sites on the PTPN22 and VDR genes respectively. We genotyped 50 patients and 50 control subjects from Lahore by using sequencing and Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (RFLP) techniques. It was observed that the allelic frequency of r and b were higher in patients as compared to controls and significantly associated with the onset of T1D. The genotype distribution frequencies varied significantly among patients and controls (p < 0.01). The mutation on the rs2476601 polymorphic site led to the change of Isoleucine to tryptophan in patients. In conclusion, compelling evidence was found of T1D onset association with the polymorphism at RsaI on rs2476601 and BsmI on rs1544410 on PTPN22 and VDR genes.