We report herein the design, synthesis, and transfection biology of a novel series of non-glycerol-based cationic lipids with lactic acid-derived head groups The synthetic procedure adopted herein for preparing 1-hydroxy-prop-2-yl head-group-based monocationic transfection lipids 1-7 is fairly straightforward and potentially applicable in designing other cationic lipids with lactic acid-derived head groups. A striking anchor-length dependency was observed in NIH3T3 cells in the sense that except lipid 4, all the other lipids were essentially transfection-inefficient. Ethidium bromide assay for the lipid:DNA interactions is consistent with the general observation that significant lipid:DNA interactions do not guarantee on improved transfection efficiency cationic lipid mediated gene delivery. Given its remarkable transfection properties and low cellular toxicity, lipid 4 is likely to find future use in the area of liposomal gene delivery.