Diversity in TLR genes coding for Toll-like receptors present in the Czech Simmental (Czech Red Pied) cattle population was tested for association with the female reproductive traits along with the traditionally assigned traits of infection resistance. The starting bull set of 164 animals represented complete gene pool. Hybrid resequencing with Illumina X-Ten WGS and PacBio amplicon sequencing yielded 7 and 9 SNPs in the TLR4 and TLR5 genes, respectively. The SNPs were validated with primer extension assays and characterized by the assignment to haplotypes and by function prediction. The breeding values for the reproductive traits, namely incidence of cystic ovaries, early reproductive disorders (ERD), calving ease, maternal calving ease, production longevity and calf vitality index were used for the association study.