Use of Reverse Genetics for the Generation of Recombinant Influenza Viruses Carrying Nanoluciferase
© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature..
Influenza A (FLUAV) and influenza B (FLUBV) viruses are human and/or animal pathogens widely studied due to their importance to public health and animal production. Both FLUAV and FLUBV possess a genome composed of eight viral gene segments. For reverse genetics of influenza viruses, transcription of the mRNA for the viral proteins is typically done from a plasmid encoding an RNA polymerase II (pol II) promoter element upstream of cloned viral cDNA and expressed like host mRNA. On the other side, the synthesis of the negative-sense, single-stranded, uncapped vRNAs can be accomplished by the host's RNA polymerase I (pol I). The reverse genetics for influenza has allowed the manipulation of influenza genomes incorporating heterogeneous sequences into different segments of the influenza genome, such as reporter genes. In this chapter, we outline the protocol from the generation of reverse genetic plasmid that can be applied for the cloning of any of the segments of FLUAV or FLUBV. Furthermore, we describe a protocol for generating FLUAV or FLUBV recombinant viruses carrying Nanoluciferase (NLuc) in the PB1 gene using reverse genetics. Finally, we delineate a microneutralization protocol using FLUAV-NLuc or FLUBV-NLuc viruses optimized for the use of antibodies from different sources (mice, ferrets, avian, etc.), which provides a more sensitive, reliable, and avidity-independent method to assess the presence of neutralizing antibodies against FLUAV or FLUBV.
| Medienart: |
E-Artikel |
|---|
| Erscheinungsjahr: |
2024 2023 |
|---|---|
| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:2733 |
|---|---|
| Enthalten in: |
Methods in molecular biology (Clifton, N.J.) - 2733(2023) vom: 08., Seite 47-74 |
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Caceres, C Joaquin [VerfasserIn] |
|---|
| Links: |
|---|
| Themen: |
Influenza |
|---|
| Anmerkungen: |
Date Revised 08.12.2023 published: Print Citation Status In-Process |
|---|
| doi: |
10.1007/978-1-0716-3533-9_4 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
NLM365549088 |
|---|
| LEADER | 01000caa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLM365549088 | ||
| 003 | DE-627 | ||
| 005 | 20231229123027.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 231226s2024 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.1007/978-1-0716-3533-9_4 |2 doi | |
| 028 | 5 | 2 | |a pubmed24n1227.xml |
| 035 | |a (DE-627)NLM365549088 | ||
| 035 | |a (NLM)38064026 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Caceres, C Joaquin |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Use of Reverse Genetics for the Generation of Recombinant Influenza Viruses Carrying Nanoluciferase |
| 264 | 1 | |c 2024 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Revised 08.12.2023 | ||
| 500 | |a published: Print | ||
| 500 | |a Citation Status In-Process | ||
| 520 | |a © 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature. | ||
| 520 | |a Influenza A (FLUAV) and influenza B (FLUBV) viruses are human and/or animal pathogens widely studied due to their importance to public health and animal production. Both FLUAV and FLUBV possess a genome composed of eight viral gene segments. For reverse genetics of influenza viruses, transcription of the mRNA for the viral proteins is typically done from a plasmid encoding an RNA polymerase II (pol II) promoter element upstream of cloned viral cDNA and expressed like host mRNA. On the other side, the synthesis of the negative-sense, single-stranded, uncapped vRNAs can be accomplished by the host's RNA polymerase I (pol I). The reverse genetics for influenza has allowed the manipulation of influenza genomes incorporating heterogeneous sequences into different segments of the influenza genome, such as reporter genes. In this chapter, we outline the protocol from the generation of reverse genetic plasmid that can be applied for the cloning of any of the segments of FLUAV or FLUBV. Furthermore, we describe a protocol for generating FLUAV or FLUBV recombinant viruses carrying Nanoluciferase (NLuc) in the PB1 gene using reverse genetics. Finally, we delineate a microneutralization protocol using FLUAV-NLuc or FLUBV-NLuc viruses optimized for the use of antibodies from different sources (mice, ferrets, avian, etc.), which provides a more sensitive, reliable, and avidity-independent method to assess the presence of neutralizing antibodies against FLUAV or FLUBV | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Influenza | |
| 650 | 4 | |a Microneutralization assays | |
| 650 | 4 | |a NanoLuc | |
| 650 | 4 | |a Reverse genetics | |
| 700 | 1 | |a Gay, L Claire |e verfasserin |4 aut | |
| 700 | 1 | |a Faccin, Flavio Cargnin |e verfasserin |4 aut | |
| 700 | 1 | |a Pérez, Daniel R |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t Methods in molecular biology (Clifton, N.J.) |d 1984 |g 2733(2023) vom: 08., Seite 47-74 |w (DE-627)NLM074849794 |x 1940-6029 |7 nnns |
| 773 | 1 | 8 | |g volume:2733 |g year:2023 |g day:08 |g pages:47-74 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1007/978-1-0716-3533-9_4 |3 Volltext |
| 912 | |a GBV_USEFLAG_A | ||
| 912 | |a GBV_NLM | ||
| 951 | |a AR | ||
| 952 | |d 2733 |j 2023 |b 08 |h 47-74 | ||