Real-time, MinION-based, amplicon sequencing for lineage typing of infectious bronchitis virus from upper respiratory samples

Salman L. Butt; Eric C. Erwood; Jian Zhang; Holly S. Sellers; Kelsey Young; Kevin K. Lahmers; James B. Stanton

The advent of real-time, in-house, third generation sequencing represents a transformative opportunity for diagnostic laboratories, by offering the ability to more fully characterize PCR reactions beyond confirming amplicon size (e.g., routine electrophoresis), Sanger sequencing RT-PCR products, or by confirming a partial sequence through probe hybridization (e.g., probe-based qPCR). However, the interpretation of such large data sets represents a challenge to veterinary diagnosticians. Read-based classification software such as genotypes from single sample. Thus, the MinION-based AmpSeq coupled with the data analysis workflow for identification, differentiation, and accurate prediction of IBV genotypes from clinical swab samples can be used as an adjunct to other established rapid diagnostic assays 9,12 until extensive testing of this protocol is done to improve and validate AmpSeq for IBV diagnostics. Furthermore, AmpSeq-based assays can be, and are being applied to other viral pathogens, 3,37 demonstrating the power and utility of this method in the age of molecular diagnostics. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/634600 doi: bioRxiv preprint