Fig. 3

Determining the allele frequency of c.151-5 T > G variant and expression of RAB3GAP1. a a tetra-primer ARMS-PCR was used to determine the allele frequency of c.151-5 T > G variant on the RAB3GAP1 gene. In this figure, well 1: heterozygous for wild-type allele, well 2: Homozygous for c.151-5 T > G variant, well 3: homozygous for the wild-type allele. b a schematic figure shows the 24 exons of RAB3GAP1 and also the effects of c.151-5 T > G on splicing. The c.151-5 T > G inserts 4 nucleotides that in turn cause frameshifting and generating the truncated protein. c Schematic of the RAB3GAP1 protein showing the Rab3 GAP domain (red) and the conserved N-terminal region of RAB3GAP1 (NTD; green). d RT-PCR was performed on RNA samples of patients and carriers (Parents). Chromatograms revealing nucleotide sequences of c.151-5 T > G of RAB3GAP1. In this figure, the size of the sequenced cDNA region is shown using 2% gel electrophoresis. e and f Relative expression levels of RAB3GAP1 in peripheral blood cells of the patient (proband), parents, and normal individual (III.3) shows a significant reduction in the patient’s mRNA level. III.3 prescreened for c.151-5 T > G variant. The experiment was performed in triplicate and ALB (albumin) was used as an internal reference gene. The data are presented as the mean ± standard deviation (SD) and the statistical significance was evaluated by Student’s t-test. Statistical analysis was performed by SPSS version 24.0 (SPSS Inc., Chicago, IL, USA) and GraphPad Prism v.8.0 (GraphPad, San Diego, USA). P- values < 0.05 were considered to be statistically significant. ** P < 0.01, *** P < 0.001