Abstract: The influence of light quality and carbon dioxide concentration on the development of photosynthetic functional structures of strawberry leaves in vitro was examined. We assessed the photochemical quenching parameter of chlorophyll a photochemical efficiency of photo system II (PSII) of strawberry leaves in vitro in a factorial set up. The main effects of light quality; averaged over CO₂ enriched, air flow and closed systems, increased the initial chlorophyll fluorescence value from 485 for yellow light developed PS II system of leaves to 1142 for white light (control) developed ones. The photochemical efficiency of PSII significantly increased from 0.64 under white light to 0.80 for yellow light developed leaves. The leaves developed under blue light were similar to that of control white light for many chlorophyll fluorescence parameters except the initial chlorophyll fluorescence level. The increase in photochemical efficiency of PSII of strawberry leaves can be attributed to lower initial fluorescence values. Under blue light the total dry weight and total chlorophyll content were increased. The possible role of high mercury peak of white light and photoinhibition during development in vitro is discussed.