Seqman software

For the sake of validation about all the exons’ sequencing consequences and segregations’ analytical results, Sanger sequencing was operated by 3500DX Genetic Analyzer (Applied Biosystems) to verify the candidates for the two families with the point mutations, while quantitative PCR (qPCR) was performed on the family with exons deletion of cDNA using the QuantStudio 5 (Applied Biosystems). With normalized reference sequences and visualizations, the sequence analysis was performed by Seqman software (Technelysium, South Brisbane, QLD, Australia). Based on the online databank of the University of California, Santa Cruz (UCSC), we designed the reference sequences. Table 1 lists the four individual primer pairs concepted for verifying the three mutations.

To verify the whole exome sequence results and segregation analysis, Sanger sequencing was performed to identify the candidate variants for the two patients and their parents using the 3500DX Genetic Analyzer (Applied Biosystems). The Seqman software (Technelysium, South Brisbane, QLD, Australia) was used for the sequence analysis with standard reference sequences and visualizations. The reference sequences were obtained using the online database at the University of California, Santa Cruz (UCSC). Two distinct primer pairs were designed and primer sequences for validating the two detected mutations are listed in Table 1.