The Utility of Deep RNAseq in Mendelian Disorder Diagnostics

To understand the importance of sequencing depth in clinical RNA-seq, we sequenced four CATS, i.e., blood, fibroblast, lymphoblastoid cells and induced pluripotent stem cells at a depth of more than 1 billion reads. We first measured the contribution of increasing sequencing depth to the detection of genes and isoforms, especially tissue-specific genes. We then present case examples whose molecular diagnoses are achieved through deep RNA-seq. We also exploit deep-RNA-seq data to provide resources for genetic diagnoses. The minimum required depth for gene and junction detection was calculated to inform the future design of clinical RNA-seq, i.e., how deep we should sequence. A splicing variation database was constructed using deep RNA-seq data to predict the splicing consequences of pathogenic variants. In conclusion, we demonstrated the utility of deep RNA-seq as a direct diagnostic tool and a database to inform the genetic diagnosis of new cases.