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. 2013 Apr 23;8(4):e61394. doi: 10.1371/journal.pone.0061394

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© 2013 Kurabi et al

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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Panel A: Ecrg4 PCR product amplified from uninfected rat ME mucosa cDNAs. A 100 bp molecular size marker is shown on the left lane. Panel B: Organization of the rat Ecrg4 gene shows the 4 exons and the start (set to +1) and stop codons of the open reading frame (ORF). The mRNA isolated from rat ME mucosa was subjected to 5′RACE analysis using a primer annealing to exon 4 (solid arrow). Following primer elongation, cloning and sequencing of the 5′ RACE products (clear bars), translation start sites (TSS) at −42 bp and −21 bp upstream of the ATG translation start site produce the identical ORF. An asterisk denotes that the −21 bp TSS was the most frequently amplified. Panel C: Immunolocalization of Ecrg4 (red) in the ME epithelium compared to a background immunofluorescence signal using pre-immune IgY (insert). Nuclei are counterstained with DAPI (blue). Epith. = epithelial cells (D) Immunoblotting of rat ME mucosa revealed the presence of a prominent 14 kDa Ecrg4 protein band.