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Fig. 4 | BMC Medical Genetics

Fig. 4

From: Segregation between SMCHD1 mutation, D4Z4 hypomethylation and Facio-Scapulo-Humeral Dystrophy: a case report

Fig. 4

DUX4 expression and characterization of the SMCHD1 mutation by RT-PCR and western blotting. a Expression of the DUX4 gene in total RNA from muscle (M) and primary fibroblasts (F) of the proband (II1). A positive control expressing DUX4-fl was used (+) and amplification was performed without reverse transcriptase (-). The ß2microglobulin gene was used as a standard of amplification. b Analysis of the SMCHD1 transcript on cDNA obtained from II1 PBMCs, muscle biopsy and primary fibroblasts. The wild-type and transcript carrying the (r.4614_4615 insTATAATA) insertion have been amplified using primers encompassing exons 36-38. PBMCs, muscle or primary fibroblasts from healthy individual (CT) were used as controls. The XNP gene was used as a positive control. c SMCHD1 western blot on whole cell extracts from PBMCs and fibroblasts from the index case (II1) compared to control cells from healthy donors (CT5 and CT7 PBMC; CT1, CT2 and CT3 primary fibroblasts) with antibodies against either the N- or C-terminal epitope. The lamin B2 protein was used as loading reference. d Primary fibroblasts were treated for 2 h with a final concentration of 50 μM NMDi14 or mock treated with DMSO. SMCHD1 transcripts were amplified by RT-QPCR in the different conditions. The ATF3 gene was used as a positive control [25]. Samples were amplified in triplicates

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