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. Author manuscript; available in PMC: 2021 Mar 1.
Published in final edited form as: Biomaterials. 2019 Dec 30;233:119750. doi: 10.1016/j.biomaterials.2019.119750

Fig. 3. Characterizations of CTB-Ang-(1–7) marker-free plants and clinical grade production.

Fig. 3.

(A) T0 seeds of plant lines #29 and 57 were grown on ½ MS0 medium without spectinomycin. After few weeks, genomic DNA was evaluated by PCR using the three sets of primers. Lane #1 WT, untransformed wild type; lanes # 2 to 6 plant # 29, Lanes # 7 to 11 plant #57; Lane# 12 positive control; lane # 13 PCR mix with H20; Lane # 14 M; 1 kb ladder (B) gDNA digested with HindIII. Lane 1, 1 kb ladder DNA; Lane 2, UT, untransformed; Lanes 3–7 DNA from marker-free seedlings of T0 plant#29; Lane 8–12 DNA from marker-free seedlings of T0 plant#57; Lane 13 T1 plant contacting the aadA gene. The position of the expected sizes of fragment is untransformed 9.1-kb, selectable marker-free is 10.7-kb, with selectable marker is 12.7-kb are indicated with arrows. Western blot of CTB-Ang-(1–7) in lettuce plants with or without the selectable marker gene (C). (D) The immunoblot assay of CTB-Ang-(1–7) expressed lettuce lyophilized leaves harvested after 30, 46 days or 60 days of plants grown in the greenhouse. (E) The immunoblot assay of CTB-Ang-(1–7) expressed in lettuce lyophilized leaves harvested at 47 days, 66 days from the plants grown in hydroponic system at Fraunhofer. 0.4 μg, 0.6μg,0.8μg of TLP Per lane was loaded and probed with anti-CTB rabbit polyclonal antibody. (F) Determination of fresh weight, dry weight, Ang1–7 expression and yield per plant of different ages grown at the Greenhouse or Fraunhofer. Fresh and dry weight per plant are calculated by normalization of total fresh and dry weight to total number of plants grown in Greenhouse (60) and Fraunhofer (800). The percentage of water content (the ratio of dry material / fresh material) are shown on the top of column.