Journal: eLife

Article Title: Gain-of-function variants in the ion channel gene TRPM3 underlie a spectrum of neurodevelopmental disorders

doi: 10.7554/eLife.81032

Figure Lengend Snippet: HEK293T cells were transfected with TRPM3 or its mutant G1007S, or a 1:1 ratio of mutant and wild-type TRPM3 plus the calcium sensor GCaMP6. Fluorescence was measured in a 96-well plate reader (Flexstation-3). ( A–C ) Representative fluorescence traces of wild-type TRPM3 ( A ), the G1007S mutant ( B ), co-transfection of TRPM3 and G1007S (1:1 ratio) ( C ). The applications of various concentrations of PS and ionomycin (Iono; 2 µM) are indicated by the arrows. Basal fluorescence was subtracted and PS-induced Ca 2+ signals were normalized to the signal after applying ionomycin. Each single trace shows the average of four replicates from the same transfection. ( D ) Hill 1 fits of the PS dose dependency of the TRPM3 mutant G1007S. Symbols represent Mean ± SEM from 3 independent experiments. For these experiments, the isoform corresponding to NCBI reference sequence NM_001366141.2 cloned in the pcDNA3.1(+)-N-eGFP vector, was used.

Article Snippet: Intracellular Ca 2+ levels were measured by a Flexstation-3 96‐well plate reader (Molecular Devices).

Techniques: Transfection, Mutagenesis, Fluorescence, Cotransfection, Sequencing, Clone Assay, Plasmid Preparation