Dental caries remains the most common chronic childhood disease. Despite strong evidence of genetic components, there have been few studies of candidate genes and caries. In this analysis we tried to assess genetic and environmental factors contributing to childhood caries in the Iowa Fluoride Study. Environmental factors (age, sex, race, tooth-brushing frequencies and water fluoride level) and three dental caries scores (d(2)fs-total, d(2)fs-pit/fissure, and d(2)fs-smooth surface) were assessed in 575 unrelated children (mean age 5.2 years). Regression analyses were applied to assess environmental correlates. The Family-Based Association Test was used to test genetic associations for 23 single nucleotide polymorphism (SNP) markers in 7 caries candidate genes on 333 Caucasian parent-child trios. We evaluated the associations between caries status and the level of both single and multiple SNPs (haplotype) respectively. Permutation procedure was performed for correction of inflated type I errors due to multiple testing. Age, tooth-brushing frequency and water fluoride level were significantly correlated to at least one carious score. Caries on pit and fissure surfaces was substantially higher than on smooth surfaces (61 vs. 39%). SNPs in three genes (DSPP, KLK4 and AQP5) showed consistent associations with protection against caries. Of note, KLK4 and AQP5 were also highlighted by subsequent haplotype analysis. Our results support the concept that genes can modify the susceptibility of caries in children. Replication analysis in independent cohorts is highly needed in order to verify the validity of our findings.