Fig. 2

In vivo and in vitro validation of the RaPrnp vector. (a) A LacZ/EGFP dual-reporter cassette was cloned into the RaPrnp vector via In-Fusion cloning. Schematic of the RaPrnp-LacZ/EGFP construct following NotI digestion. (b) Brightfield, (c) EGFP fluorescence signal, and (d) merge of brains from 2-month-old Tg12084 rats. Left: Tg; right: non-Tg. Rat (e–g) and mouse (h–j) cells were transfected with the RaPrnp-LacZ/EGFP construct. Neuronal rat (e and f) and mouse (h and i) cells were transfected along with rat (g) and mouse (j) fibroblasts. EGFP fluorescence was detected in B35 (e), PC12 (f), N2a (h), and CAD5 (i) neuronal cell lines. A few transfected RAT2 fibroblast cells had low EGFP fluorescence (g). Mouse 3T3 cells transfected with the RaPrnp-LacZ/EGFP vector did not support EGFP fluorescence (j). Arrowheads = cytoplasm; arrows = neurite-like extensions