Aims: Flocculent wine yeasts were characterized for the expression of FLO1, FLO5, FLO8, AMN1 and RGA1 genes, growth kinetics and physicochemical properties of the cell surface during a 6-month sparkling wine fermentation period. Methods and Results: The expression of FLO1, FLO5, FLO8, AMN1 and RGA1 genes was determined by RT-qPCR. The physicochemical characterization of yeast surface properties was evaluated by the microbial adhesion to solvents method. FLO5 gene was the most expressed one and a linear correlation with the flocculent degree was found. Flocculent strains were more hydrophobic than the commercial wine strain EC1118. Conclusions: Gene expressions and the ability to face secondary wine fermentation conditions were strain dependent. The importance of FLO5 gene in developing the high flocculent characteristic of wine yeasts was highlighted. Cell surface properties depended on the time of fermentation. Significance and Impact of the Study: Better knowledge about the expression of some genes encoding the flocculent phenotype which could be useful to select suitable starter cultures to improve sparkling wine technology was achieved. A step forward in understanding the complexity and strain-specific nature of flocculation phenotype was done.