Abstract:  The aim of this study is to define a reliable reckoning of gene frequencies and six‐locus haplotypic frequencies of HLA‐A, HLA‐B, HLA‐C, HLA‐DRB1, HLA‐DQA1, HLA‐DQB1 and HLA‐DPB1 in the Tunisian population. One hundred unrelated random, healthy people originating from various parts of Tunisia were typed for the alleles of the loci mentioned above by using the molecular techniques polymerase chain reaction – hybridization with oligonucleotide probe (PCR‐SSO) and sequence specific primers (SSP). The population studied appeared to be in Hardy–Weinberg equilibrium. Allelic frequency distributions were observed at each locus. The most frequent HLA‐A alleles were HLA‐A*02 (39%) HLA‐A*0101 (25%), HLA‐A*30 (21%) and HLA‐A*2301 (18%). Moreover, HLA‐3A*3601, HLA‐1A*6601, HLA‐1A*3402 and HLA‐2A*8001 were found; however, no HLA‐A*4301 was detected. For the HLA‐B locus, the most common in descending order were HLA‐B*44 (22%), HLA‐B*5001 (19%), HLA‐B*51 (16%) and HLA‐B*18 (15%). Among the 28 alleles HLA‐Cw detected, HLA‐Cw*6 and HLA‐Cw*7 were highly predominant with the frequencies of 33 and 30%, respectively. For the HLA class‐II loci, HLA‐DRB1*0701, HLA‐DRB1*11, HLA‐DRB1*13 and HLA‐DRB1*03 were the most frequent DR alleles. For the HLA‐DPB1, HLA‐DPB1*0401, HLA‐DPB1*0301 and HLA‐DPB1*0201 were the most frequent DP alleles. Many haplotypes were in a strong positive‐linkage disequilibrium. The most frequent haplotypes for HLA‐A, HLA‐B, HLA‐C and HLA‐DRDQ were HLA‐A*3301, HLA‐B*1402, HLA‐Cw*0802, HLA‐DRB1*0102, HLA‐DQA1*0101 and HLA‐DQB1*0501; HLA‐A*2402, HLA‐B*0801, HLA‐Cw*0702, HLA‐DRB1*0301, HLA‐DQA1*0501 and HLA‐DQB1*0201; HLA‐A*2902, HLA‐B*4403.1, HLA‐Cw*1601, HLA‐DRB1*0701, HLA‐DQA1*0201 and HLA‐DQB1*0202; HLA‐A*3002, HLA‐B*1801, HLA‐Cw*0501, HLA‐DRB1*0301, HLA‐DQA1*0501 and HLA‐DQB1*0201, with frequencies between 0.025 and 0.015. These data can be used as control data for HLA disease associations and paternity studies, but they are also important for the evaluation of the probability rate of success in determining the optimal matched donor in unrelated stem transplantation for Tunisian patients or patients of Tunisian origin.