There are diverse types of gelling agents that are used in media cultures. Agar and Gelrite are among the gelling agents used in DKW culture medium, as the common culture medium for the micropropagation of walnut plants. The effects of these gelling agents have not been investigated on the successful production ofin vitroexplants and the response of the explants to ex vitro evaporative demand is overlooked so far. Stomata and water relations of tissue culture medium determine the successful production ofin vitroplants, therefore this experiment was conducted to investigate the effect of two types of gelling agents (Agar and Gelrite) on the stomatal characteristics, transpiration rate (E), and desiccation responses ofin vitrowalnut explants. Stomatal morphology, transpiration rate, RWC, and some morpho-physiological traits such as shoot length, chlorophyll content, osmotic potential (ψs), proline, and glycine betaine content were evaluated in micropropagated walnut explants cultured on Agar or Gelrite. Analysis of results indicated no considerable changes in the morpho-physiological characteristics of explants grown in DKW medium containing Agar or Gelrite gelling agents. Compared with the medium containing Agar, adding Gelrite to the DKW medium caused a decrease in E and an increase in relative leaf water content (RWC) of the walnut explant's leaves during desiccation. Gelrite induced generation of more closed stomata leading to a reduction in E and increase in RWC during desiccation. This resulted in improvement of walnut plantlet's capacity to conserve their water content and as the consequence promoted ability to prevent ex vitro wilting. Manuscript profile