A one-step real-time reverse transcription-polymerase chain reaction (RT-PCR) using a TaqMan probe to quantitatively detect spring viremia of carp virus (SVCV) is described. In this assay, a pair of primers amplifying an 81-bp DNA fragment and a TaqMan probe was designed targeting the conserved region at the SVCV glycoprotein (G) gene. To avoid the disadvantages arising from plasmids, an extension adding a T7 phage polymerase promoter to the 5’ end of the antisense primer was carried out to obtain viral cRNA. Standardized cycle threshold (Ct) values for 10-fold serial dilutions of SVCV cRNA were achieved by real-time RT-PCR and used to create standard curves. A regression line between the mean Ct values and viral template concentrations over a 1:10⁷ dilution range with an r² value (0.9916) and a slope (-3.36) and the coefficient of variation (intra- or inter-assay is <2% and <4%, respectively) indicated that the assay was highly reproducible. The assay was specific to SVCV and there was no cross-reactivity with other fish viruses (viral hemorrhagic septicemia virus, VHSV infectious pancreatic necrosis virus, IPNV grass carp reovirus, GCRV epizootic haematopoietic necrosis virus, EHNV). The standard curve allows precise absolute quantitation and shows that the detection limit of the assay is 40 copies of the viral RNA. This one-step RT-PCR assay was evaluated using 60 clinical common carp samples collected during the year 2006, indicating such technology offers considerable advantages over conventional RT-PCR methods in current routine use for SVCV surveillance. This is the first report of the development of a one-step TaqMan^® RT-PCR for SVCV detection. Journal of Virological Methods 152 (2008) 43–48. Zhiqin Yuea,, Yong Teng, Chengzhu Liang, Xiayang Xie, Biao Xua, Laihua Zhu,Zhiwen Lei, Junqiang He, Zongxiao Liu, Yulin Jiang, Hong Liu, Qiwei Qin.( Shandong Entry-Exit Inspection and Quarantine Bureau, No. 1 Tuandao Er Road, Qingdao 266002, China State Key Laboratory of Biocontrol, College of Life Sciences, SunYat-Sen (Zhongshan) University, 135 West Xingang Road, Guangzhou 510275, China The Key Lab of Aquatic Animal Diseases, Shenzhen Exit & Entry Inspection and Quarantine Bureau, 2049 Heping Road, Shenzhen 518001, China Medical College of Georgia, Augusta, GA 30912, USA).