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Fig. 4 | BMC Biology

Fig. 4

From: A de novo silencer causes elimination of MITF-M expression and profound hearing loss in pigs

Fig. 4

A new generated silencer in the M-promoter eliminated Mitf-m transcription. a Transcriptional activity analysis of the Mitf-m promoter from the R and r alleles. The reporter constructs are shown on the left, and the corresponding relative luciferase activity measured in transient transfection assays is shown on the right. The luciferase activity of pGL3-r-7.8 k was significantly lower than that of pGL3-R-7.8 k. There was no significant difference between the R and r alleles when constructs were shorter than 7.8 k. Error bars indicate the standard deviations. The results shown are for one experiment with four technical replicates. Raw data for this and three additional experiments with similar results are provided in Additional file 5; sheet 5, Data of reporter assay. b Schematics of the M-promoter. The sequence variations between the R and r alleles are labeled. INS, insertion; DEL, deletion. c Schematics of the M-promoter from –7513 bp to –7609 bp relative to the transcription start site of the M-exon. Sequence differences between R and r allele are indicated with a red box. The new sites showing consensus sequence for SOX protein binding are underlined in red. The oligonucleotide probes designed for electrophoretic mobility shift assay (EMSA) are highlighted. d EMSA shows the specific binding of the nuclear proteins to the r2 probe, and absence of binding to the R2 probe. In vitro incubation was performed using the indicated nuclear extracts, probe and unlabeled oligonucleotides (cold probes). C1, complex 1; C2, complex 2; R, R2 probe; r, r2 probe; N, random (negative control) probe

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