Control cells (wt/wt) and the homozygous cell line from the Finnish patient (W262X/W262X) were treated with 100 μg/ml cycloheximide (CHX) for 3 hours

<p><b>Copyright information:</b></p><p>Taken from "A minor alternative transcript of the fumarylacetoacetate hydrolase gene produces a protein despite being likely subjected to nonsense-mediated mRNA decay"</p><p>BMC Molecular Biology 2005;6():1-1.</p><p>Published online 7 Jan 2005</p><p>PMCID:PMC546004.</p><p></p> Total RNA was extracted and subjected to RT-PCR. RAR serves as a control for RNA quantity in each sample. Control amplifications (showing an increasing number of cycles) for del100, del231 and RAR are displayed on the left to show that, with the conditions used, each PCR reaction is in the exponential phase. (A) [αP]-dATP was incorporated during the PCR reaction and the products loaded on a 6% acrylamide gel. The gel was used to directly expose an X-ray film and the signal was quantified using the NIH Image 1.2 software. FAH was amplified from exons 6 to 14 (the del100 transcript which is detected using these primers is indicated by a star). Del100 and del231 were amplified using the specific primers RT84 and RT85. (B) Quantification of the amount of del100 mRNA in 3 different experiments, normalized to RAR levels. The error bars represent standard deviations. Del100 is stabilized between 4- to 7-fold in homozygous cells (W262X/W262X) and normal cells (wt/wt) following the cycloheximide treatment (yellow bars: - cycloheximide; purple bars: + cycloheximide). (C) del231 is relatively unaffected by translation inhibition