Figure 1. Generation of ORMDL3 TG mice.

(A). The human ORMDL3 transgenic construct contains a CAG promoter for universal over-expression (light blue), H2B-mRFP (red fluorescent protein)(red), followed by a transcriptional stop site (orange), the human ORMDL3 open reading frame (green), and a rabbit B-globin poly-adenylation sequence (pink). The H2B-mRFP (red fluorescent protein) and transcriptional stop site are flanked by LoxP sites (triangles). Cells containing this pCAGEN Lox mRFP-H2B STOP Lox hORMDL3 construct will express RFP (red fluorescent protein) and not express hORMDL3 as the transcriptional stop codon (orange) prevents transcription of hORMDL-3. (B). Expression of the hORMDL3 gene is Cre recombinase dependent since the transcriptional stop codon preventing hORMDL-3 expression (and RFP) are excised by Cre recombinase via the Lox P sites resulting in over-expression of hORMDL3 only in those cells expressing Cre recombinase. (C) Primer sets (F1, R; F2, R) used to detect progeny of RFP-Stop^FLhORMDL3-TG mice crossed with Zp3-cre mouse and predicted sizes of transgene mRNA assessed by PCR. (D) WT mice (left) and hORMDL3^zp3-Cre mice (right) aged 26 weeks appeared morphologically similar. (E) The gross appearance of lungs of WT mice (left) and hORMDL3^zp3-Cre mice (right) aged 26 weeks appeared morphologically similar. (F). Other than airway remodeling the appearance of lungs of WT mice (left) and hORMDL3^zp3-Cre mice (right) aged 26 weeks appeared morphologically similar. (G) Expression of the human ORMDL3 (hORMDL3) transgene was detected by qRT/PCR in mouse lung, (H) mouse bronchial epithelium, and (I) mouse BAL macrophages in hORMDL3^zp3-Cre mice, but not in WT mice. The human ORMDL3 transgene did not increase levels of mouse ORMDL1 (m-ORMDL1), mouse ORMDL2 (m-ORMDL2), or mouse ORMDL3 (m-ORMDL3) as assessed by qRT/PCR in mouse lung (J), mouse bronchial epithelium (K), and mouse BAL macrophages (L) in either hORMDL3^zp3-Cre or WT mice.