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Updated: May 17, 2026

Using Reverse Genetics to Manipulate the NSs Gene of the Rift Valley Fever Virus MP-12 Strain to Improve Vaccine Safety and Efficacy
Published on: November 1, 2011
Murine hepatitis virus nsp4 N258T mutants are not temperature-sensitive
Dia C Beachboard1, Xiaotao Lu, Susan C Baker
1Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, 1161 21st Avenue South, Nashville, TN 37232, USA.
Abstract:
Coronavirus replicase nsp4 is critical for virus-induced membrane modifications. An nsp4 mutant (N258T) of murine hepatitis virus (MHV) has been reported to be temperature-sensitive (ts) and to alter membrane targeting. We engineered and recovered all four possible codon variants of N258T in the cloned MHV-A59 background. All mutant viruses demonstrated impaired replication compared to wildtype MHV, but no nsp4 N258T mutant virus was ts, and all variants colocalized with viral protein markers for replication complexes, but not with markers for mitochondria. This study emphasizes that complete genome sequencing may be necessary, even with directed and confirmed reverse genetic mutants.

