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Research Article
Szu-Cheng Kuo
National Defense Medical Center
Corresponding Author
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Reverse genetics is an important tool in the elucidation of viral replication and the development of countermeasures; however, these methods are impeded by laborious and inefficient replicon delivery methods. This paper demonstrates the use of a baculovirus to facilitate the efficient delivery of autonomous CHIKV replicons into mosquito and mammalian cells in vitro as well as adult mosquitoes in vivo. The efficacy of this approach was verified via co-localization among an eGFP reporter, nsP1, and dsRNA as well as through the inhibition of an RNA-dependent RNA polymerase (RdRp) null mutation (DDAA) in nsP4, or the treatment of a known antiviral compound (6-azauridine). We also investigated the correlation between CHIKV replicon-launched eGFP expression and the effectiveness of CHIKV replicon variants in inducing IFN-β expression in human cell lines. This delivery method based on a single vector is applicable to mosquito and mammalian cells in seeking to decipher the mechanisms underlying CHIKV replication, elucidate virus–host interactions, and develop antivirals. This study presents an effective alternative to overcome many of the technological issues related to the study and utilization of autonomous arbovirus replicons.
Keywords
Reverse genetics, development, CHIKV, DDAA
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No competing interests reported.
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On 22 Mar, 2021
Reviewer #10 agreed
On 22 Mar, 2021
Reviewer #6 agreed
On 22 Mar, 2021
Reviewer #1 agreed
On 22 Mar, 2021
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On 22 Mar, 2021
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This preprint is under consideration at Scientific Reports. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research Article
Szu-Cheng Kuo
National Defense Medical Center
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Review
Version 1
Posted 03 Mar, 2021
No community comments so far
Reviewer #11 agreed
On 22 Mar, 2021
Reviewer #2 agreed
On 22 Mar, 2021
Reviewer #9 agreed
On 22 Mar, 2021
Reviewer #7 agreed
On 22 Mar, 2021
Reviewer #10 agreed
On 22 Mar, 2021
Reviewer #6 agreed
On 22 Mar, 2021
Reviewer #1 agreed
On 22 Mar, 2021
Reviewer #5 agreed
On 22 Mar, 2021
Reviewers invited
Invitations sent on 01 Mar, 2021
Editor assigned
On 01 Mar, 2021
Editor invited
On 23 Feb, 2021
Submission checks complete
On 23 Feb, 2021
First submitted
On 18 Feb, 2021
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Reverse genetics is an important tool in the elucidation of viral replication and the development of countermeasures; however, these methods are impeded by laborious and inefficient replicon delivery methods. This paper demonstrates the use of a baculovirus to facilitate the efficient delivery of autonomous CHIKV replicons into mosquito and mammalian cells in vitro as well as adult mosquitoes in vivo. The efficacy of this approach was verified via co-localization among an eGFP reporter, nsP1, and dsRNA as well as through the inhibition of an RNA-dependent RNA polymerase (RdRp) null mutation (DDAA) in nsP4, or the treatment of a known antiviral compound (6-azauridine). We also investigated the correlation between CHIKV replicon-launched eGFP expression and the effectiveness of CHIKV replicon variants in inducing IFN-β expression in human cell lines. This delivery method based on a single vector is applicable to mosquito and mammalian cells in seeking to decipher the mechanisms underlying CHIKV replication, elucidate virus–host interactions, and develop antivirals. This study presents an effective alternative to overcome many of the technological issues related to the study and utilization of autonomous arbovirus replicons.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8
Figure 9
Figure 10
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