Viruses constitute a major constraint to soybean production worldwide and are responsible for significant yield losses every year. Although varying degrees of resistance to specific viral strains has been identified in some soybean genetic sources, the high rate of mutation in viral genomes and mixed infections of different viruses or strains under field conditions usually hinder the effective control of viral diseases. In the present study, we generated transgenic soybean lines constitutively expressing the double-strand RNA specific ribonuclease gene PAC1 from Schizosaccharomyces pombe to evaluate their resistance responses to multiple soybean-infecting virus strains and isolates. Resistance evaluation over three consecutive years showed that the transgenic lines displayed significantly lower levels of disease severity in field conditions when challenged with soybean mosaic virus (SMV) SC3, a prevalent SMV strain in soybean-growing regions of China, compared to the non-transformed (NT) plants. After inoculation with four additional SMV strains (SC7, SC15, SC18, and SMV-R), and three isolates of bean common mosaic virus (BCMV), watermelon mosaic virus (WMV), and bean pod mottle virus (BPMV), the transgenic plants exhibited less severe symptoms and enhanced resistance to virus infections relative to NT plants. Consistent with these results, the accumulation of each virus isolate was significantly inhibited in transgenic plants as confirmed by quantitative real-time PCR and double antibody sandwich enzyme-linked immunosorbent assays. Collectively, our results showed that overexpression of PAC1 can increase multiple virus resistance in transgenic soybean, and thus provide an efficient control strategy against RNA viruses such as SMV, BCMV, WMV, and BPMV.