Sendai virus (SeV) is a non-segment negative-sense RNA virus that naturally infects and causes pneumonia in mice. As a prototypic member of the family Paramyxoviridae, SeV has been characterized well, and these studies revealed numerous traits of paramyxovirus biology. The reverse genetics system to rescue SeV was first established in 1995. The virus was rescued from a cloned cDNA that contains full genome sequence flanked by T7 promoter and hepatitis delta virus ribozyme. To rescue SeV, it is necessary to infect cells with a recombinant vaccinia virus vTF7.3 that expresses T7 RNA polymerase, and transfect with the SeV full genome cDNAs together with supporting plasmids encoding NP, P, and L genes under the T7 promoter. Synthesized viral RNA by T7 RNA polymerase will be encapsidated with NP and associated with a polymerase complex composed of P and L. The polymerase complex transcribes and replicates the genome, and produces progeny virions. Rescued SeV needs to be plaque purified to exclude vTF7.3 from viral stock. Reverse genetics system of SeV is relatively efficient compared to other paramyxoviruses, but alternative approaches to rescue poorly growing mutant viruses are also available.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others ReferencesEgelman EH, Wu SS, Amrein M, Portner A, Murti G (1989) The Sendai virus nucleocapsid exists in at least four different helical states. J Virol 63(5):2233–2243
Calain P, Roux L (1993) The rule of six, a basic feature for efficient replication of Sendai virus defective interfering RNA. J Virol 67(8):4822–4830
Kato A, Sakai Y, Shioda T, Kondo T, Nakanishi M, Nagai Y (1996) Initiation of Sendai virus multiplication from transfected cDNA or RNA with negative or positive sense. Genes Cells 1(6):569–579
Garcin D, Pelet T, Calain P, Roux L, Curran J, Kolakofsky D (1995) A highly recombinogenic system for the recovery of infectious Sendai paramyxovirus from cDNA: generation of a novel copy-back nondefective interfering virus. EMBO J 14(24):6087–6094
Leyrer S, Neubert WJ, Sedlmeier R (1998) Rapid and efficient recovery of Sendai virus from cDNA: factors influencing recombinant virus rescue. J Virol Methods 75(1):47–58
Sutter G, Ohlmann M, Erfle V (1995) Non-replicating vaccinia vector efficiently expresses bacteriophage T7 RNA polymerase. FEBS Lett 371(1):9–12
Ghanem A, Kern A, Conzelmann KK (2012) Significantly improved rescue of rabies virus from cDNA plasmids. Eur J Cell Biol 91(1):10–16. doi:10.1016/j.ejcb.2011.01.008
Maddon PJ, Dalgleish AG, McDougal JS, Clapham PR, Weiss RA, Axel R (1986) The T4 gene encodes the AIDS virus receptor and is expressed in the immune system and the brain. Cell 47(3):333–348
Fuerst TR, Niles EG, Studier FW, Moss B (1986) Eukaryotic transient-expression system based on recombinant vaccinia virus that synthesizes bacteriophage T7 RNA polymerase. Proc Natl Acad Sci U S A 83(21):8122–8126
Zhan X, Slobod KS, Krishnamurthy S, Luque LE, Takimoto T, Jones B, Surman S, Russell CJ, Portner A, Hurwitz JL (2008) Sendai virus recombinant vaccine expressing hPIV-3 HN or F elicits protective immunity and combines with a second recombinant to prevent hPIV-1, hPIV-3 and RSV infections. Vaccine 26(27–28):3480–3488
Takahashi T, Ryan KW, Portner A (1992) A plasmid that improves the efficiency of foreign gene expression by intracellular T7 RNA polymerase. Genet Anal Tech Appl 9(3):91–95
Takimoto T, Bousse T, Portner A (2000) Molecular cloning and expression of human parainfluenza virus type 1 L gene. Virus Res 70(1–2):45–53
Bousse T, Matrosovich T, Portner A, Kato A, Nagai Y, Takimoto T (2002) The long noncoding region of the human parainfluenza virus type 1 f gene contributes to the read-through transcription at the m-f gene junction. J Virol 76(16):8244–8251
Kovacs GR, Vasilakis N, Moss B (2001) Regulation of viral intermediate gene expression by the vaccinia virus B1 protein kinase. J Virol 75(9):4048–4055. doi:10.1128/JVI.75.9.4048-4055.2001
Wyatt LS, Moss B, Rozenblatt S (1995) Replication-deficient vaccinia virus encoding bacteriophage T7 RNA polymerase for transient gene expression in mammalian cells. Virology 210(1):202–205. doi:10.1006/viro.1995.1332
Sakai Y, Kiyotani K, Fukumura M, Asakawa M, Kato A, Shioda T, Yoshida T, Tanaka A, Hasegawa M, Nagai Y (1999) Accommodation of foreign genes into the Sendai virus genome: sizes of inserted genes and viral replication. FEBS Lett 456(2):221–226
Li HO, Zhu YF, Asakawa M, Kuma H, Hirata T, Ueda Y, Lee YS, Fukumura M, Iida A, Kato A, Nagai Y, Hasegawa M (2000) A cytoplasmic RNA vector derived from nontransmissible Sendai virus with efficient gene transfer and expression. J Virol 74(14):6564–6569
Tsung K, Yim JH, Marti W, Buller RM, Norton JA (1996) Gene expression and cytopathic effect of vaccinia virus inactivated by psoralen and long-wave UV light. J Virol 70(1):165–171
Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, Rochester, NY, 14642, USA
Shringkhala Bajimaya, Tsuyoshi Hayashi & Toru Takimoto
Correspondence to Toru Takimoto .
Editor information Editors and AffiliationsDepartment of Population Health, Poultry Diagnostic and Research Center, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
Daniel R. Perez
© 2017 Springer Science+Business Media LLC
About this protocol Cite this protocolBajimaya, S., Hayashi, T., Takimoto, T. (2017). Rescue of Sendai Virus from Cloned cDNA. In: Perez, D. (eds) Reverse Genetics of RNA Viruses. Methods in Molecular Biology, vol 1602. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6964-7_7
Download citationDOI: https://doi.org/10.1007/978-1-4939-6964-7_7
Published: 16 May 2017
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-6962-3
Online ISBN: 978-1-4939-6964-7
eBook Packages: Springer Protocols
RetroSearch is an open source project built by @garambo | Open a GitHub Issue
Search and Browse the WWW like it's 1997 | Search results from DuckDuckGo
HTML:
3.2
| Encoding:
UTF-8
| Version:
0.7.4