Cloning and Sequencing of Phytoplasma Ribosomal DNA (rONA) Associated with Kerala Wilt Disease of Coconut Palms

Using the polymerase chain reaction (PCR) the 165 rRNA gene of phytoplasma associated with Kerala wilt disease of coconut palm (Cocos nucifera L) was amplified from infected leaf samples. Within the three universal primer pairs P1/P6, P1/P7and P41 P7, the primer pair P4/P7 only showed an amplification of 650 bp DNA fragment. 5ince P4/P7 amplifies the 16S-23S intergenic spacer region of 165 rRNA gene, the PCR product 650 bp of Kerala wilt disease palm indicates the phytoplasma DNA. The amplified fragment was sequenced and deposited in Genbank data library (Accession No. AY158660). The absence of restriction sites for Bcll and Rsa/l in 650 bp indicates phytoplasmic nature of DNA and its strain difference. A comparison of the 650 bp sequence with other phytoplasmas and its restriction profile indicates Kerala wilt disease phytoplasma as a separate 165 rRNA group in the classification of phytoplasmas. To our knowledge, this report records the first finding of the phytoplasma DNA using universal primers and its sequence analysis in coconut palms of Kerala, south India.