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Flavonol synthase (FLS) is a key enzyme for the formation of flavonols, which are a subclass of the flavonoids. FLS catalyses the conversion of dihydroflavonols to flavonols. The enzyme belongs to the 2-oxoglutarate-dependent dioxygenases (2-ODD) superfamily. We characterized the<em>FLS</em>gene family of<em>Brassica napus</em>that covers 13 genes, based on the genome sequence of the<em>B. napus</em>cultivar Express 617. The goal was to unravel which<em>BnaFLS</em>genes are relevant for seed flavonol accumulation in the amphidiploid species<em>B. napus.</em>Two<em>BnaFLS1</em>homoelogs were identified and shown to encode bifunctional enzymes. Both exhibit FLS activity as well as flavanone 3-hydroxylase (F3H) activity, which was demonstrated<em>in vivo</em>and<em>in planta. BnaFLS1-1</em>and -<em>2</em>are capable of converting flavanones into dihydroflavonols and further into flavonols. Analysis of spatio-temporal transcription patterns revealed similar expression profiles of<em>BnaFLS1</em>genes. Both are mainly expressed in reproductive organs and co-expressed with the genes encoding early steps of flavonoid biosynthesis. Our results provide novel insights into flavonol biosynthesis in<em>B. napus</em>and contribute information for breeding targets with the aim to modify the flavonol content in rapeseed.