Ultra-rapid somatic variant detection via real-time targeted amplicon sequencing.
Commun Biol
; 5(1): 708, 2022 07 15.
Article
in En
| MEDLINE
| ID: mdl-35840782
ABSTRACT
Molecular markers are essential for cancer diagnosis, clinical trial enrollment, and some surgical decision making, motivating ultra-rapid, intraoperative variant detection. Sequencing-based detection is considered the gold standard approach, but typically takes hours to perform due to time-consuming DNA extraction, targeted amplification, and library preparation times. In this work, we present a proof-of-principle approach for sub-1 hour targeted variant detection using real-time DNA sequencers. By modifying existing protocols, optimizing for diagnostic time-to-result, we demonstrate confirmation of a hot-spot mutation from tumor tissue in ~52 minutes. To further reduce time, we explore rapid, targeted Loop-mediated Isothermal Amplification (LAMP) and design a bioinformatics tool-LAMPrey-to process sequenced LAMP product. LAMPrey's concatemer aware alignment algorithm is designed to maximize recovery of diagnostically relevant information leading to a more rapid detection versus standard read alignment approaches. Using LAMPrey, we demonstrate confirmation of a hot-spot mutation (250x support) from tumor tissue in less than 30 minutes.
Full text:
1
Database:
MEDLINE
Main subject:
Neoplasms
Type of study:
Diagnostic_studies
/
Guideline
/
Prognostic_studies
Limits:
Humans
Language:
En
Journal:
Commun Biol
Year:
2022
Type:
Article
Affiliation country:
United States