Jembrana disease is an acute viral disease  in Bali cattle that have short incubation period, high mortality and morbidity rate. Early diagnostic methods are needed  to prevent the further spreading of this disease. In this study, we combined One-step RT-PCR  and  NALF  methods to detect env-tm gene of Jembrana virus Tabanan 1987 strain. Viral RNA isolated from a spleen of the infected cattle was used as the template. One-step RT-PCR procedure was performed using oligoprobes  labeled digoxigenin and  right primer sequence that were designed using the primer3plus program based  on the conserves region of env-tm gene from NCBI database. The product of one-step RT-PCR was tested using NALF  method  instead of  electrophoresis. Positive result was shown by the appearance of dark lines on the test line of digoxigenin in NALF dipstick device.