To investigate whether human leukocyte antigen-G (HLA-G) gene polymorphism is associated with in vitro fertilization (IVF) failure, we sequenced exons 2-4 of the HLA-G gene in 50 couples with three or more IVFs (including 10 couples with five or more IVFs) and 58 control fertile couples from a Polish population. Of the 10 different HLA-G alleles identified in our study subjects, neither allele was found to be associated with IVF. We also genotyped 50 couples with IVF and 71 control couples for the -725C>G variant in the promoter region and the 14 bp insertion or deletion polymorphism in the 3' untranslated region of the HLA-G gene. The frequency of -725GG or GC genotype in women with IVF and in control fertile women was similar [26% vs 25.3%; odds ratio (OR) = 1.0; P = 1.0]. The 14 bp ins/ins or ins/del genotype was more common in women with IVF than in control women (76.9% vs 59.1%; OR 2.4; P = 0.03), but the difference was not significant after Bonferroni correction for multiple comparisons. The frequency of the ins/ins or ins/del genotype was particularly high (90%) in women who experienced five or more IVFs (OR = 6.2; P = 0.08), but again, the excess was not statistically significant, possibly because of small sample sizes. These results are in line with functional studies that show lower levels of HLA-G mRNA and protein related to the HLA-G allele including the 14 bp sequence and suggest that the insertion allele may be associated with an increased risk of IVF.