Fluorine configuration at C2' of the bis(2'-fluorothymidine) dinucleotide is demonstrated to drive intramolecular base stacking. 2'-β F-Configuration drastically reduces stacking compared to the 2'-α series. Hence, base stacking emerges as being tunable by the C2'-F stereoconfiguration through dramatic puckering variations scrutinized by NMR and natural bond orbital analysis. Accordingly, 2'-β F-isomer photoreactivity is significantly reduced compared to that of the 2'-α F-isomer.