The tomato plant exhibiting leaf rolling, witches’ broom, distorted and elongated flower’s sepals inIğdır province, Turkey, was observed. Total DNA extraction was performed from the symtomaticfresh tomato sample. All DNAs were subjected to Direct and Nested polymerase chain reaction (PCR)with universal primer sets that amplified the 16S rRNA of phytoplasmas. PCR products were purifiedfrom agarose gel and cloned into the pGEM T-Easy cloning vector. Recombinant plasmids wereintroduced into the prokaryotic cloning bacteria by electroporation. Plasmid isolation was performedby selecting one of the positive clones randomly and sequencing was performed by Next GenerationSequencing (NGS). Sequencing results revealed that the 16S rRNA gene associated with phytoplasmawas 1251 nucleotides in size, and this sequence was denominated as ‘Iğdır 10’ isolates and recordedin the GenBank under the MT344968 accession number. The virtual restriction fragment lengthpolymorphism (V-RFLP) and phylogenetic analysis of the 16S rDNA sequence confirmed that thecause of disease in infected tomato plants was 'Candidatus Phytoplasma trifolii' ('Ca. P. trifolii')(16SrVI-A, Clover proliferation group), with a 1.00 similarity coefficient. This present study is thefirst report of ‘Ca. P. trifolii’ and its nucleotide sequence analysis in naturally infected tomato in Iğdırprovince.