Rapid Sequencing of Multiple RNA Viruses in their Native Form

Wongsurawat, T.; Jenjaroenpun, P.; Taylor, M. K.; Lee, J.; Tolardo, A. L.; Parvathareddy, J.; Kandel, S.; Wadley, T. D.; Kaewnapan, B.; Athipanyasilp, N.; Skidmore, A. M.; Chung, D.; Chaimayo, C.; Whitt, M.; Kantakamalakul, W.; Sutthent, R.; Horthongkham, N.; Ussery, D. W.; Jonsson, C. B.; Nookaew, I.

doi:10.1101/482471

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bioRxiv · 10.1101/482471

Rapid Sequencing of Multiple RNA Viruses in their Native Form

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Abstract

Long-read nanopore sequencing by a MinION device offers the unique possibility to directly sequence native RNA. We combined an enzymatic poly-A tailing reaction with the native RNA sequencing to (i) sequence complex population of single-stranded (ss)RNA viruses in parallel, (ii) detect genome, subgenomic mRNA/mRNA simultaneously, (iii) detect a complex transcriptomic architecture without the need for assembly, (iv) enable real-time detection. Using this protocol, positive-ssRNA, negative-ssRNA, with/without a poly(A)-tail, segmented/non-segmented genomes were mixed and sequenced in parallel. Mapping of the generated sequences on the reference genomes showed 100% length recovery with up to 97 % identity. This work provides a proof of principle and the validity of this strategy, opening up a wide range of applications to study RNA viruses.

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Wongsurawat, T., Jenjaroenpun, P., Taylor, M. K., Lee, J., Tolardo, A. L., Parvathareddy, J., Kandel, S., Wadley, T. D., Kaewnapan, B., Athipanyasilp, N., Skidmore, A. M., Chung, D., Chaimayo, C., Whitt, M., Kantakamalakul, W., Sutthent, R., Horthongkham, N., Ussery, D. W., Jonsson, C. B., Nookaew, I.. 2018-11-29. Rapid Sequencing of Multiple RNA Viruses in their Native Form. https://doi.org/10.1101/482471

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