We have examined the prothrombin gene as a candidate gene for venous thrombosis in selected patients with a documented familial history of venous thrombophilia. All the exons and the 5′- and 3′-UT region of the prothrombin gene were analyzed by polymerase chain reaction and direct sequencing in 28 probands. Except for known polymorphic sites, no deviations were found in the coding regions and the 5′-UT region. Only one nucleotide change (a G to A transition) at position 20210 was identified in the sequence of the 3′-UT region. Eighteen percent of the patients had the 20210 AG genotype, as compared with 1% of a group of healthy controls (100 subjects). In a population-based case-control study, the 20210 A allele was identified as a common allele (allele frequency, 1.2%; 95% confidence interval, 0.5% to 1.8%), which increased the risk of venous thrombosis almost threefold odds ratio, 2.8; 95% confidence interval, 1.4 to 5.6. The risk of thrombosis increased for all ages and both sexes. An association was found between the presence of the 20210 A allele and elevated prothrombin levels. Most individuals (87%) with the 20210 A allele are in the highest quartile of plasma prothrombin levels (> 1.15 U/mL). Elevated prothrombin itself also was found to be a risk factor for venous thrombosis.