Rapid sample preparation method for high molecular weight DNA from tumor tissues suitable for structural variant analysis

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Molecular & Cellular Analysis Technologies
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Not Applicable
SummaryClaremont BioSolutions, LLC (CBIO)The introduction of long-read sequencing and mapping applications is set to change the field of cancerresearch by allowing unparalleled insight into the complex structure of tumor genomes. With the ability tosequence large spans of DNA that are 10s-100s of kb in size these technologies will offer more accurate denovo assembly, resolve highly repetitive regions, and allow identification of large structural variants in tumorgenomes. As cancer researchers move to these platforms they will face new hurdles as effective samplepreparation of ultra-long DNA (>250 kb) remains a limiting step. Current methods of DNA extraction, includingspin column and magnetic bead applications, significantly shear ultra-long DNA. Older, albeit effective,agarose plug digestion techniques are difficult, time-consuming and highly variable. New commercial productshave been introduced that can isolate ultra-long DNA but are restricted in use to bacteria, tissue culture cells,or fresh blood cells, and are not usable with clinically relevant solid tumor tissue samples. To overcome thischallenge, Claremont BioSolutions is developing a prototype technology that combines gentle and rapid tissuedissociation with a novel binding matrix to isolate high quality/high quantity ultra-long DNA in <20 minutes. Themethod reduces DNA shearing and tangling and is compatible with small quantities of solid tissue. In Aim 1, weshall use the combined approach to demonstrate isolation of ultra-long DNA (>250 kb) from matched normaland tumor tissue samples and analyze length and quality. In Aim 2, we shall validate compatibility of theisolated ultra-long DNA with third generation long-read sequencing and mapping technologies and analyze thesequence data to demonstrate improved long-read sequencing over existing DNA extraction methods.Ultimately, the goal will be to integrate this miniature technology into an automated platform to provide a rapid,cost effective, and reproducible method to isolate DNA from solid tissues for improved downstream analysis oftumor samples.