By comparing three expression vectors for the rabies virus (Rv) minigenome, we show that the characteristic of the Rv RNA is important for efficient rescue despite its not being crucial for replication. Moreover, we show that the coexpression of the viral proteins from helper Rv and Mokola virus could rescue the Rv minigenome while Rv-related European bat lyssavirus 1 could not, suggesting that...

To study the replication of Nipah virus (NiV), a minigenome replication assay that does not require the use of infectious virus was developed. The minigenome was constructed to encode a NiV vRNA analogue containing the gene for chloramphenicol acetyltransferase (CAT) under the control of putative NiV transcription motifs and flanked by the NiV genomic termini. CAT protein was detected only when...

(2015) Establishment of a minigenome system for oropouche virus reveals the S genome segment to be significantly longer than reported previously, Abstract 33 34 Oropouche virus (OROV) is a medically important orthobunyavirus, which causes 35 frequent outbreaks of a febrile illness in the Northern parts of Brazil. However, despite 36 being the cause for an estimated half a million human infectio...

The role of the variable portion of the noncoding regions (NCRs) of the three Bunyaviridae RNA segments (L, M, S) in transcription, replication, and packaging was studied using the recently developed plasmid-driven RNA polymerase I minigenome system for Uukuniemi (UUK) virus, genus Phlebovirus (11), as a model. Comparison of the different segments showed that all NCRs were sufficient to mediate...

BACKGROUND Embryonated chicken eggs have been used since the mid-20th century to grow a wide range of animal viruses to high titers. However, eggs have found so far only limited use in the production of recombinant proteins. We now describe a system, based on a Sendai virus minigenome, to produce large amounts of heterologous viral glycoproteins in the allantoic cavity of embryonated eggs. RE...

The genome of lymphocytic choriomeningitis virus (LCMV) consists of two negative-sense single-stranded RNA segments, designated L and S. Both segments contain two viral genes in an ambisense coding strategy, with the genes being separated by an intergenic region (IGR). We have developed a reverse genetic system that allows the investigation of cis-acting signals and trans-acting factors involve...

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne virus (genus Nairovirus, family Bunyaviridae) associated with high case fatality disease outbreaks in regions of Africa, Europe, and Asia. The CCHFV genome consists of three negative-strand RNA segments, S, M, and L. The unusually large virus L polymerase protein and the need for biosafety level 4 (BSL-4) containment conditions for w...

During the construction of the transmissible gastroenteritis virus (TGEV) full-length cDNA clone, a point mutation at position 637 that was present in the defective minigenome DI-C was maintained as a genetic marker. Sequence analysis of the recovered viruses showed a reversion at this position to the original virus sequence. The effect of point mutations at nucleotide 637 was analyzed by rever...

We developed a system to identify the viral proteins required for the packaging and passage of human respiratory syncytial virus (RSV) by reconstructing these events with cDNA-encoded components. Plasmids encoding individual RSV proteins, each under the control of a T7 promoter, were cotransfected in various combinations together with a plasmid containing a minigenome into cells infected with a...

We describe an Ebolavirus minigenome-based system that is suitable for high-throughput screening of compounds able to impair Ebolavirus virus replication and/or transcription. The assay is robust (Z' factor, >0.6) and can be carried out in low-biosafety containment. Results from a pilot screen of 960 compounds are presented.