Phytoplasmas as one of the most important plant pathogens cause serious damage to grape around the world. Recently, symptoms such as leaf curl, yellowing and purpling were observed in central region of Iran resembled those caused by phytoplasma diseases. To identify and grouping grape infecting phytoplasmas in central region of Iran, some samples showing leaf curl, yellowing and purpling were collected. DNA was extracted from leaf veins of collected samples and used as template in PCR. PCR method was carried out for phaytoplasma detection using 16S rRNA region universal primers. Universal Primer pairs P1/P7 and P1A/P17 were used in direct PCR. In PCR the expected size of 1784 bp DNA fragment was amplified in a few samples using primer pair P1/P7 but, most samples showed the expected size of 1759 bp using primer pair P1A/P17. Nested-PCR was performed using primer pair R16F2n/R16R2 to amplify the internal regions of the 16S-23S rRNA. This primer pair amplified a 1239 bp DNA fragment in most samples collected in late summer and early fall but not in the spring and early summer samples. Nested-PCR was also performed using primer pairs fUu5/rU3, PA2F/R and NPA2F/R. Primer pairs FUu5/rU3, PA2F/R and NPA2F/R amplified 876 bp, 1178 bp and 485 bp DNA fragments in positive samples, respectively. The PCR results revealed primer pair P1A/P7A for direct PCR and primer pair R16F2n/R16R2 for nested-PCR were the best primes for grape phytoplasmas detection. Also, primer pairs FUu5/rU3, PA2F/R and NPA2F/R were found suitable for this purpose. Grape phytoplasmas were characterized by PCR-RFLP. The amplified 1239 bp DNA fragment of different isolates was digested by Hea III, Hinf I, Mse I, Msp I ( Hap II), Rsa I and Taq I restricted enzymes. PCR-RFLP analysis showed that the isolates are justify; LINE-HEIGHT: normal; MARGIN: 0in 0in 0pt" Key words: Grape, Ca. Phytoplasma solani Stolbor group (16SrXII), PCR-RFLP analysis, Sequencing
