Begin the preparation of the glass slide by cutting a rectangular piece of clear, double-sided nano tape and sticking it to the center of a clean glass slide without removing the protective plastic film on the top side of the tape. Next, dissect the chute by removing the upper portion of the older leaves with a wire stripper. Then hold the chute by the mesocaudal and carefully remove the surrounding leaves with a dental probe to extract the primordia of interest.
For mounting the primordium, remove the protective film from the tape. Lay the exposed primordium on the tape. Cut the primordium at the base using a razor blade, discarding the basal stem and the hypocaudal.
For smoother unrolling, lubricate the inner, or, adaxial leaf surface by dipping the needle tip into 100%glycerol. Unroll the primordium using a dental probe with a bent needle. Position the needle tip parallel to the surface and unfurl the outer basal margin by gently pressing it against the tape.
With the outer margin adhering to the tape, align the needle tip parallel to the long axis of the leaf. Gently slide the needle sideways to unroll and flatten the leaf onto the tape. Apply a drop of water to the unrolled primordium.
Immediately place a cover slip on the water drop and the primordium. Gently press down the edges of the cover slip so that they adhere to the tape. Once done, place the slide on the epifluorescence microscope stage to visualize the fluorophore.
This method was followed to visualize and analyze the fluorescent protein expression in maize leaf primordia. Complementary to the transverse section analysis. The whole leaf mount analysis revealed tissue and stage-specific patterns of formal response during vein formation.
Whole mount analysis method enabled qualitative and quantitative analyses of fluorescent protein expression in the maize leaf primordia, indicating the effectiveness of this method in examining the maize leaf primordia which are difficult to image due to their rolled morphology.
