AbstractAims  Protein kinase C (PKC), a serine/threonine kinase, is known to be activated in various tissues under hyperglycaemic conditions. Notably, PKCβ, a member of the conventional PKC group, is the predominant isoform detected in vascular tissues and could be involved in the development of diabetic vascular complications. In the present study, we investigated genetic variations in the 5′‐upstream region of the PKCβ gene to assess their possible relation to vascular complications in diabetic patients.Methods  Variations upstream from the PKCβ gene (−1066/+256) were examined in 60 Type 2 diabetic patients using a cycle sequencing method. Screening of detected variations was performed in 204 Type 2 diabetic patients and 160 healthy controls.Results  Five single nucleotide polymorphisms; C(−238)G, C(−287)T, A(−348)G, C(−546)G, and C(−853)T, were identified in the upstream region. The C(−287)T and A(−348)G polymorphisms were in perfect linkage disequilibrium. There were no significant differences in genotype or allele frequencies of the five polymorphisms among the diabetic patients and healthy subjects. However, both −238GG and −287CC (−348GG) homozygotes showed significantly higher frequencies of macrovascular disease compared with patients with other genotypes. Further, an electrophoretic mobility shift assay revealed that the −238G fragment had a five‐fold higher affinity for transcription factor Sp1 when compared with −238C.Conclusions  The C(−238)G and C(−287)T‐A(−348)G polymorphisms in the 5′‐upstream region of the PKCβ gene may have an effect on the susceptibility of diabetic vascular complications through an alteration of tissue PKCβ density or function.