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. 2016 Aug 5;11(8):e0160385. doi: 10.1371/journal.pone.0160385

Fig 2. Amplification, cloning and translation of ERG isoforms.

Fig 2

A) Analysis of PCR-amplified ERG isoforms by electrophoresis on agarose gel. PCR products with length corresponding to ERG3 coding region (1458bp) were present in all samples. Calculated length of amplified coding region of predicted aberrant ERG isoforms was 775bp in NALM6 and ALL samples 1,2,4 and 5 (bearing the deletion of ERG exons 7–13) and 880bp in ALL sample 3 (bearing the deletion of ERG exons 7–11). PCR products corresponding to predicted aberrant ERG isoforms were present in all ERGdel-positive samples and in none ERGdel-negative samples (B-other cases: ALL-6 and ALL-7, hyperdiploid cases: ALL-8 and ALL-9, ETV6/RUNX1-positive case—ALL-10). (B) Schematic representation of ERG transcript variants cloned from NALM6 and of predicted encoded proteins. Regions encoded by alternative frames are displayed in grey. (C) Analysis of ERGaber proteins synthesized from PCR-prepared DNA templates by in vitro transcription/translation (T/T) assay. Proteins were detected by western blot using following antibodies: Anti-ERG antibody EPR3863 (Ab-N), Erg-1/2/3 Antibody C-17 (Ab-int), Erg-1/2/3 Antibody C-20 (Ab-C). In vitro T/T reaction without any DNA template served as a negative control (Neg. Ctrl.). ERGaberN was detected by Ab-N only and ERGaberC by Ab-C only. Ab-int showed only unspecific binding to proteins present in reticulocyte lysate used for T/T assay.