RESUMEN
Genome sequencing (GS) is a powerful clinical tool used for the comprehensive diagnosis of germline disorders. GS library preparation typically involves mechanical DNA fragmentation, end repair, and bead-based library size selection followed by adapter ligation, which can require a large amount of input genomic DNA. Tagmentation using bead-linked transposomes can simplify the library preparation process and reduce the DNA input requirement. Here we describe the clinical validation of tagmentation-based PCR-free GS as a clinical test for rare germline disorders. Compared with the Genome-in-a-Bottle Consortium benchmark variant sets, GS had a recall >99.7% and a precision of 99.8% for single nucleotide variants and small insertion-deletions. GS also exhibited 100% sensitivity for clinically reported sequence variants and the copy number variants examined. Furthermore, GS detected mitochondrial sequence variants above 5% heteroplasmy and showed reliable detection of disease-relevant repeat expansions and SMN1 homozygous loss. Our results indicate that while lowering DNA input requirements and reducing library preparation time, GS enables uniform coverage across the genome as well as robust detection of various types of genetic alterations. With the advantage of comprehensive profiling of multiple types of genetic alterations, GS is positioned as an ideal first-tier diagnostic test for germline disorders.
Asunto(s)
ADN , Enfermedades Raras , Humanos , Secuencia de Bases , Mapeo Cromosómico , Análisis de Secuencia de ADN/métodos , Biblioteca de Genes , Secuenciación de Nucleótidos de Alto Rendimiento/métodosRESUMEN
INTRODUCTION: Genotype-phenotype discordance occurs occasionally in congenital adrenal hyperplasia (CAH). Its causes are largely unknown. We describe a case of monochorionic, diamniotic twins with discordant clinical presentations of CAH, and show evidence for this being due to mosaicism resulting from a postzygotic full gene deletion of CYP21A2 prior to twinning. CASE DESCRIPTION: A 7-day-old 36-week gestation female infant (Twin A) presented to the emergency department with elevated 17-hydroxyprogesterone (17-OHP). Her identical twin (Twin B) had normal 17-OHP on newborn screening. Both twins showed signs of virilization, more pronounced in Twin B. Molecular genetic testing of both twins and their parents showed a WT paternally-inherited CYP21A2 and a maternally-inherited copy containing the c.293-13C>G mutation. Both twins were also found to have a 5'-CYP21A1P/CYP21A2-3' hybrid (product of the common 30-kb deletion), derived from the deletion of the paternally-inherited CYP21A2. Neither mother nor father carried the deletion. CONCLUSIONS: The genetic findings are consistent with mosaicism for two CYP21A2 cell lines in both twins. The first cell line is expected, based on parental results, while the second line is due to a postzygotic full gene deletion of the paternally-inherited WT CYP21A2. The resultant genotype, compound heterozygosity for c.293-13C>G and a CYP21A2 full gene deletion, is consistent with a salt-wasting CAH phenotype. Differential distribution of the second cell line between the twins is most likely the cause for their discrepant phenotypes. We believe this is the first report of somatic CYP21A2 mosaicism, and represents a novel cause for discrepant CAH phenotypes in monozygotic twins.