The transcriptome analysis of tartary buckwheat identified a flavonol synthase gene FtFLS1. In order to further understand its structure， function and diversity in tartary buckwheat genome， we identified 104 members of FLS gene family with 10 subgroups， in which FtFLS1 was found in DF8 subgroup. Promoter analysis revealed two MeJA response elements at the upstream of 1500 bp sequence. We analyzed the expression of FtFLS1 in different organs and its response to MeJA treatments. The transcriptional level of FtFLS1 in stems and leaves was comparable but higher in roots. Expression of FtFLS1 also increased significantly with the treatment of MeJA. We subsequently cloned the CDS sequence of FtFLS1， and then generated FtFLS1 over-expressed hairy root lines of tartary buckwheat and detected their flavonoid content. Over-expression transformants over accumulated the downstream products of FLS， which including kaempferol， quercetin and rutin， while the contents of dihydrokaempferol and dihydroquercetin， the substrates of brass synthetase， decreased significantly. Furthermore， we analyzed the diversity of FtFLS1 gene in different populations of tartary buckwheat， and found that Northern landraces， Southwestern landraces and Himalayan wild accessions present obvious differentiation. The results are helpful for understanding the FtFLS1-mediated synthesis of flavonoids and the domestication process of buckwheat.