Abstract

Identification of Clinically Significant Genetic Variants by Exome Sequencing of DNA Extracted from Dried Blood Spots: A Model for Newborn Screening

Toumy Guettouche, Director, Children’s Hospital of Philadelphia

Current analytical methods for newborn screening are based on detection of small metabolites for the diagnosis of inborn errors of metabolism (IEM) to prevent long- term complications. However an analytical gap exists as there is an increasing number of genetic disorders that lack the analytical screening technique that is needed for reliable detection. Genomic methods allow early identification of clinically relevant genetic variants and hold the promise to augment or replace currently used new born screening techniques and reduce the analytical gap. Whole exome sequencing (WES) is a primary method for screening of genomic variants but standard methods require significant amounts of DNA for successful analysis. The most common method for storage of newborn patient blood samples are dried blood spots. However, a major challenge using dried blood spot samples for genetic studies is the small amount of DNA (typically less than 300ng/3mm punch) that can be extracted which is significantly below the input required for standard WES protocols. Here we present a protocol that allows WES from as little as 75ng of genomic DNA extracted from a dried blood spot. We sequenced the exomes of 4 patients with previously identified lysosomal storage disorders. The sequencing results generated by our method are comparable to results obtained by standard WES protocols and allowed identification of the pathogenic variant and confirmation of the diagnosis in these patients. All steps of our protocol are amenable to automation and therefore adaptable to high-throughput processing. To our knowledge this is the first report of the successful identification of a genetic variant by whole exome sequencing from DNA extracted from a dried blood spot.