Abstract: It is one of the important control strategies of rice orange leaf disease by development of a rapid disease detection technique. Conserved general primer pairs sP1 and sP2 for phytoplasma were synthesized, and the 558 bp segment of the phytop!asma 16S rRNA gene was specifically amplified by PCR. The sequencing results of the segment showed that the nucleotide acid sequence of this PCR product shared above 95% identical with the corresponding segment of other kinds of phytoplasma in Genbank, which testified that the PCR product was from the pathogen of rice orange leaf disease. The PCR detection system was established by optimizing the parameters of PCR reaction system and found that this system could effectively, sensitively, and accurately detected the disease. The several samples from the different locations of Xinyi, Gaozhou and Conghua in Guangdong Province were detected by the PCR system, and the results revealed rice leaf disease existed in these locations.