Abstract
This chapter reports the high-throughput sequencing protocol for sequencing Coronaviruses and other positive strand viruses to produce a dataset of significant depth of coverage. The protocol describes sequencing of infectious bronchitis virus propagated in embryonated eggs and harvested in the allantoic fluid. The protocol is composed of three main steps—enrichment of the allantoic fluid using ultracentrifugation, extraction of total RNA from allantoic fluid, and library preparation from total RNA to DNA sequencing libraries. The workflow will be suitable for all coronaviruses using high-throughput sequencing platforms.
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References
Zhou P, Yang XL, Wang XG, Hu B, Zhang L, Zhang W, Si HR, Zhu Y, Li B, Huang CL, Chen HD, Chen J, Luo Y, Guo H, Jiang RD, Liu MQ, Chen Y, Shen XR, Wang X, Zheng XS, Zhao K, Chen QJ, Deng F, Liu LL, Yan B, Zhan FX, Wang YY, Xiao GF, Shi ZL (2020) A pneumonia outbreak associated with a new coronavirus of probable bat origin. Nature 579(7798):270–273. https://doi.org/10.1038/s41586-020-2012-7
Garvey MI, Bradley CW, Holden KL, Hewins P, Ngui SL, Tedder R, Jumaa P, Smit E (2017) Use of genome sequencing to identify hepatitis C virus transmission in a renal healthcare setting. J Hosp Infect 96(2):157–162. https://doi.org/10.1016/j.jhin.2017.01.002
Houlihan CF, Frampton D, Ferns RB, Raffle J, Grant P, Reidy M, Hail L, Thomson K, Mattes F, Kozlakidis Z, Pillay D, Hayward A, Nastouli E (2018) Use of whole-genome sequencing in the investigation of a nosocomial influenza virus outbreak. J Infect Dis 218(9):1485–1489. https://doi.org/10.1093/infdis/jiy335
Andino R, Domingo E (2015) Viral quasispecies. Virology 479-480C:46–51. https://doi.org/10.1016/j.virol.2015.03.022
Guy JS (2015) Isolation and propagation of coronaviruses in embryonated eggs. In: Maier HJ, Bickerton E, Britton P (eds) Coronaviruses: methods and protocols. Springer New York, New York, NY, pp 63–71. https://doi.org/10.1007/978-1-4939-2438-7_7
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Freimanis, G.L., Oade, M.S. (2020). Whole-Genome Sequencing Protocols for IBV and Other Coronaviruses Using High-Throughput Sequencing. In: Maier, H., Bickerton, E. (eds) Coronaviruses. Methods in Molecular Biology, vol 2203. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0900-2_5
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DOI: https://doi.org/10.1007/978-1-0716-0900-2_5
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0899-9
Online ISBN: 978-1-0716-0900-2
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