All patients were informed in person and their written consent was obtained. Genomic DNA was isolated from peripheral blood leucocytes using the QIAamp DNA Blood Mini QIAcube Kit (Qiagen, Hilden, Germany), according to the manufacturer’s protocols. All coding exons and exon-intron boundaries of seven genes that were associated with MODY (
KCNJ11, ABCC8, INS, GCK, HNF4A, HNF1A, HNF1B) were amplified using the Multiplicom MODY MasterDx (Agilent, CA, USA) kit. Prepared library was sequenced on the Illumina Miseq platform (Illumina Inc., San Diego, CA, USA). The data were analyzed by the Sophia DDM data analysis software. In order to call variants, sequencing data was aligned to human reference genome, hg19. After amplifying targeted regions using designed primers, Sanger sequencing on ABI Prism 3500 Genetic Analyzer (Thermo Fisher Scientific, MA USA) was performed for the confirmation of the detected variants and segregation analysis. Novel variations were classified according to the American College of Medical Genetics and Genomics criteria (
20 (link)
). Mutation Taster, The Sorting Intolerant from Tolerant (SIFT) and deleterious annotation of genetic variants using neural networks (DANN) were used for computational pathogenicity prediction (
21 (link)
). The data of minor allele frequencies of variants were obtained from GnomAD (
22 (link)
).
KCNJ11, ABCC8, INS, GCK, HNF4A, HNF1A, HNF1B) were amplified using the Multiplicom MODY MasterDx (Agilent, CA, USA) kit. Prepared library was sequenced on the Illumina Miseq platform (Illumina Inc., San Diego, CA, USA). The data were analyzed by the Sophia DDM data analysis software. In order to call variants, sequencing data was aligned to human reference genome, hg19. After amplifying targeted regions using designed primers, Sanger sequencing on ABI Prism 3500 Genetic Analyzer (Thermo Fisher Scientific, MA USA) was performed for the confirmation of the detected variants and segregation analysis. Novel variations were classified according to the American College of Medical Genetics and Genomics criteria (
20 (link)
). Mutation Taster, The Sorting Intolerant from Tolerant (SIFT) and deleterious annotation of genetic variants using neural networks (DANN) were used for computational pathogenicity prediction (
21 (link)
). The data of minor allele frequencies of variants were obtained from GnomAD (
22 (link)
).
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