African Journal of
Biotechnology

  • Abbreviation: Afr. J. Biotechnol.
  • Language: English
  • ISSN: 1684-5315
  • DOI: 10.5897/AJB
  • Start Year: 2002
  • Published Articles: 12213

Full Length Research Paper

Silencing of the rift valley fever virus s-genome segment transcripts using RNA interference in Sf21 insect cells

Juliette Rose Ongus
  • Juliette Rose Ongus
  • Jomo Kenyatta University of Agriculture and Technology, P. O. Box 62000-00200 Nairobi, Kenya.
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Evans Kiplangat Rono
  • Evans Kiplangat Rono
  • Jomo Kenyatta University of Agriculture and Technology, P. O. Box 62000-00200 Nairobi, Kenya.
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Fred Alexander Wafula Wamunyokoli
  • Fred Alexander Wafula Wamunyokoli
  • Jomo Kenyatta University of Agriculture and Technology, P. O. Box 62000-00200 Nairobi, Kenya.
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  •  Received: 23 January 2017
  •  Accepted: 10 April 2017
  •  Published: 03 May 2017

Abstract

Rift Valley fever (RVF) is a mosquito-borne disease that affects humans and livestock. It results in economically disastrous livestock deaths in Africa. The causative agent is RVF virus (RVFV), which possesses a tripartite negative-sense, single-stranded RNA genome, with large (L), medium (M) and small (S) segments. The lack of effective vaccines and anti-RVFV therapies motivates concerted efforts in search for safe and effective remedies. Recent advances in the RNA interference (RNAi) may provide promising tools to control RVFV. This study aimed to use the Baculovirus Expression Vector System to demonstrate that the RVFV S-genome segment is susceptible to RNAi. A multiplex short hairpin RNA (shRNA) transcription cassette was constructed for generating small interfering RNA (siRNA) that target both the RVFV genomic RNA transcripts and the Green Fluorescent Protein (GFP) which was used as a reporter gene. Two expression vectors were constructed; one for expressing the RVFV S-genome alone and the other one for simultaneous expression of both the RVFV S-genome and the shRNA cassette. A separate transiently expressed GFP only vector was included as an internal positive control to be expressed simultaneously when co-transfected into Sf21 insect cells with each RVFV-S construct to monitor the effectiveness of the shRNA to trigger RNAi. By design, the RNAi effect on both the RVFV-S and the GFP transcripts was driven from the same shRNA. A statistically significant reduction in the relative GFP fluorescence (P< 0.05) was demonstrated and a drop in GFP and the RVFV S transcript levels observed. Collectively, these results suggest the potential application of RNAi as an antiviral strategy against RVFV.

 

Key words: Baculovirus Expression, rift valley fever, RNA interference, green fluorescent protein.