nm 000261  (OriGene)

Journal: PLoS ONE

Article Title: Mutant myocilin impacts sarcomere ultrastructure in mouse gastrocnemius muscle

doi: 10.1371/journal.pone.0206801

Figure Lengend Snippet: ( A ) Western blots for human MYOC protein in adult transgenic mouse gastrocnemius muscle lysates. Human MYOC protein was detected in the transgenic mouse skeletal muscle and in the heart using R&D Systems anti-MYOC antibody (1:1000). Loading was 40μg tissue lysate per well of a 10% SDS-PAGE gel. Total mouse number for this representative western blot is N = 6 different animals. ( B ) NTM5 cells were transiently-transfected with a plasmid with cDNA for human MYOC-FLAG or a plasmid with cDNA for mouse Myoc-FLAG. 20μg cell lysates were treated with PNGase to remove N- glycosylation followed by Western blot analysis using anti-FLAG (1:1000). ( C ) Weights of gastrocnemius muscles and hearts harvested from equal numbers of male and female wt and CMV-MYOC-Y437H transgenic mice did not differ between groups. N per group was ≥ 6. +/- SD is indicated and t-test were p>0.1. Abbreviations–wild-type, wt; transgenic, Tg.

Article Snippet: NTM5 cells at 70 to 80% confluence were transiently-transfected with FLAG-tagged plasmids (8μg total cDNA per 10cm plate), human MYOC (Origene, RC206556; Accession number NM_000261), or mouse Myoc (Origene, MR224777; Accession NM_010865) using FuGENE6 transfection reagent (Promega, E2691).

Techniques: Western Blot, Transgenic Assay, SDS Page, Transfection, Plasmid Preparation