GFP and GUS localization in transgenic tobacco plants generated for the construct GFP::P85–P87::GUS.

<p>(A–C) Confocal laser scanning microscopic analysis of GFP expression under the <i>At4g35985</i> promoter (P85) in 21-day-old tobacco seedlings (T₂ progeny). GFP expression in leaf midrib (A), apical meristematic region (B) and primary root (C) are presented. Green fluorescence image of GFP (left); bright field image (middle), superimposed image (right) are shown. Bar represents 250 µm in each image. Histochemical GUS detection under <i>At4g35987</i> promoter (P87) in transgenic tobacco plants, GUS expression in germinating tobacco seeds (D) and young primordia at 14-day-old seedlings (E), bar represents 1 mm. (F to H) GUS expression in 21-day-old tobacco seedlings (F); GUS expression in tobacco flower (G); and reproductive tissues of tobacco flower (H); bar represents 5 mm. (I to K) GUS expression in the base of the filament of tobacco flower (I), anther (J) and stigma (K), bar represents 1 mm. (L) GUS expression in pollen grains, bar represents 100 µm. (M) Immunoblot analysis of GFP protein accumulation in various tissue of transgenic tobacco plants generated for the construct GFP::P85–P87::GUS expressing GFP under P85. Western blot showing GFP expression in young leaf, older leaf, stem, flower and root tissues of tobacco plant (T₂ generation). Forty µg of total protein from different tissues were subjected to 10% polyacrylamide gel electrophoresis, anti GFP antibody from Santa Cruz Biotechnology (Santa Cruz, CA) was used as primary antibody and horseradish peroxidase-conjugated anti-rabbit secondary antibody (1∶5000) in western blot. Lower panel represents the loading control by detecting Rubisco large subunit (LSU) using Ponceau S stain.