Transcriptome analysis of TSC1 and TSC2 using Nanopore long-read sequencer

Tuberous sclerosis complex (TSC) is a relatively common autosomal dominant disorder characterized by multiple dysplastic organ lesions and neuropsychiatric symptoms, caused by loss-of-function mutation of either TSC1 or TSC2. Target-capture full-length double-stranded cDNA sequencing using long-read sequencer Nanopore (Nanopore Long-read Target Sequencing) revealed that the various kinds of the TSC1 and TSC2 full-length transcripts and the novel intron retention transcripts of TSC2 in TSC patient. Our results indicate that the Nanopore Long-read Target Sequencing is useful for the detection of mutations and confers information on the full-length alternative splicing transcripts for the genetic diagnosis. Overall design: Total RNA extracted from the peripheral blood mononuclear cells (PBMCs) of two controls and TSC patient were used for the full-length double-stranded cDNA synthesis with SMARTer method. Then the full-length double-stranded cDNA generated by SMARTer method was captured using TSC1 and TSC2 probe and then the captured double-stranded cDNA was ligated adapter for Nanopore long-read sequencing using the ligation sequencing kit. The libraries were sequenced on Nanopore MinION long-read sequencer.