Assessment of genetic stability of long-term cultivation of plantlet derived tTCL Citrumelo using repetitive sequence primers. Regeneration of plantlet from organogenesis of stem transverse thin cell layer (tTCL) was achieved for Citrumelo, a valuable rootstock. Identification of the genetic stability of plant tissue culture is absolutely necessary. The aim of this study was to assess the potential retrotransposons and inter simple sequence repeat (ISSR) primers in detecting the genetic stability of the Citrumelo plantlet derived from tTCL technique. The research was conducted from Juni 2013 until October 2015 in Breeding Laboratory of Indonesian Citrus and Subtropical Fruits Research Institute. A four repetitive based sequences retrotransposon and ISSR marker assays were used to evaluate genetic stability of a group of 22 months old in vitro plantlets and to confirm the most reliable method for true-to-type propagation of Citrumelo. Leaves of plantlets were selected and isolated in bulk. Groups of DNA in bulk segregant analysis (BSA) were amplified and separated using agarose gel. Vitroplants that morphologically similar have been effectively distinguished by a selected primer INT- retrotransposon that detect an deletion band at 550 bp on a line a group of sample. Retrotransposon is abundance through the genome and its activation induced by stress condition. Tissue culture condition was reported potential to induce retrotransposon activation. The genetic variation of 2.6% was acceptable for the culture that produced from long-term. Plantlets used in this study were selected from population induced from early 2014, and employed for studying media as well as environment factors for efficiently organogenesis of citrumelo in period of 2014-2015. However, additional study is on going for evaluating genetic variability from a cycle plantlet production through tTCL of stems. This combination protocol of organogenesis and PCR based markers detection would be powerful tools for mass propagation of high quality seedling derived tissue culture for breeding or cultivation programs.