A new Defective Helper RNA to produce Sindbis virus that infects neurons but does not propagate

CC-BY-NC 4.0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not. Abstract Recombinant Sindbis viruses are important tools in neuroscience because they combine rapid and high transgene expression with large payloads. Currently, two packaging systems are available for making neurotropic virus, but both produce a fraction of viral particles that can propagate beyond the primary infected neuron. When injected into mouse brains, viruses produced using these systems label neurons at the injection site, but also elsewhere in the brain. Such ectopic labeling caused recombinant Sindbis viruses to be classified as anterograde viruses with limited retrograde spread, and can complicate the interpretation of neuroanatomical and other experiments. Here we describe a new packaging system, DH-BB(5'SIN;TE12ORF), that produces virus that is both neurotropic (able to infect neurons) and propagation-incompetent. We show in mice that DH-BB(5'SIN;TE12ORF)-packaged virus eliminates infection of cells outside the injection site. We also provide evidence that the ectopically labeled cells observed in previous experiments resulted from secondary infection by propagation-competent virus, rather than from inefficient retrograde spread. Virus produced with our new packaging system retains all the advantages of previous recombinant Sindbis viruses, but minimizes the risks of confounding results with unwanted ectopic labeling. It should therefore be considered in future studies in which a neurotropic, recombinant Sindbis virus is needed.. CC-BY-NC 4.0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not .