3-hour genome sequencing and targeted analysis to rapidly assess genetic risk

This proof-of-concept experiment demonstrates how prior knowledge of haplotype structure or familial variants can be used to rapidly evaluate an individual at risk for a genetic disease. While ultra-rapid sequencing remains both complex and cost prohibitive, our method is more easily automated than prior approaches and uses smaller volumes of blood, thus may be more easily adopted for future studies of ultra-rapid genome sequencing in the clinical setting.

Following extraction of DNA from cord blood and rapid library preparation, genome sequencing was performed on an Oxford Nanopore PromethION. FASTQ files were generated from original sequencing data in near real-time and aligned to a reference genome. Variant calling and analysis were performed at timed intervals.

Rapid genetic testing in the critical care setting may guide diagnostic evaluation, direct therapies, and help families and care providers make informed decisions about goals of care. We tested whether a simplified DNA extraction and library preparation process would enable us to perform ultra-rapid assessment of genetic risk for a Mendelian condition, based on information from an affected sibling, using long-read genome sequencing and targeted analysis.

We optimized the DNA extraction and library preparation methods to create sufficient library for sequencing from 500 μL of blood. Real-time, targeted analysis was performed to determine that the newborn was neither affected nor a heterozygote for variants underlying a Mendelian condition. Phasing of the target region and prior knowledge of the affected haplotypes supported our interpretation despite a low level of coverage at 3 hours of life.