Technical Abstract: Tobacco streak virus (TSV) is a pathogen that has been reported in soybean in Brazil and the United States, for which resistance has only been reported in the “hay-type” soybean cultivar Tanner. To find additional resistant soybean genotypes, over 1000 soybean accessions from the USDA Soybean Germplasm Collection were evaluated for resistance to TSV in the greenhouse. No resistance was found in a 94 accession ancestral set, but 19 of 926 accessions screened from a geographically diverse set of soybean accessions were resistant to TSV infection. Further investigation showed that the resistance of the soybean accessions was temperature-sensitive. When TSV-resistant plants were inoculated with TSV and grown at 24°C for 17-20 days in a growth chamber, only 7% of inoculated plants became infected, based on virus antigen detected in noninoculated trifoliolate leaves. In contrast, when inoculated with TSV and grown at 32°C for 17-20 days, 71% of TSV-resistant plants became systemically infected with TSV, including plants of the resistant control, Tanner, which was earlier identified as resistant to TSV infection and had never shown systemic TSV movement under greenhouse conditions. Because temperatures may equal or exceed 32°C at times during the growing season in many soybean producing areas, it will be important to assess the potential usefulness of the TSV resistance under field conditions for several years before utilizing it in breeding programs.