Engineering of a Sagiyama Alphavirus RNA‐Based Transient Expression Vector

• DOI: 10.1111/j.1348-0421.2002.tb02668.x
• Publication Date: 2013-11-14T15:35:10Z
• Authors: Yuka Yamaguchi; Yukio Shirako
• Keywords: Sindbis virus; Subgenomic mRNA

Abstract Sagiyama virus (SAGV), a strain of Getah in the genus Alphavirus family Togaviridae , has broad host range vertebrates and invertebrates but is not pathogenic for humans. We engineered SAGV genome as an efficient transient expression vector using full‐length infectious cDNA clone pSAG2 background. A green fluorescent protein (GFP) gene was used reporter expressed from subgenomic mRNA. When GFP placed downstream intact capsid or internally deleted encoding N‐terminal 9 amino acids C‐terminal 149 acids, autoproteolysis occurred efficiently at boundary site to release N‐terminally‐fused capsid‐protease domain. also without 5′‐terminal region gene, suggesting that does have translation enhancer sequence. To provide structural proteins pseudovirion formation, nonviable mutant construct, pSAG2.3L, which contains Gly‐to‐Leu substitution − 2 position nsP3/4 cleavage site, helper. up 50 μg 1 × 10 6 BHK21 cells after inoculation pseudovirions. The C6/36 mosquito cell suitable large scale In addition high‐level expression, safeness should give advantage over other alphavirus vectors.