Characterization of A Kinetochore Protein Knl2 in Solanum Lycopersicum

Background and aim: a major challenge in plant breeding programs is to create true breeding lines, for production of hybrid plants. classically these lines are produced by time-consuming rounds of inbreeding. alternatively haploid plants can be generate through the culture of haploid tissues like anther and ovule by in vitro approaches which is not yet available or is restricted to the particular number of genotypes. therefore any improvement in the haploid induction systems to increase the level of haploid plants production is of high interest. recently a promising centromere- mediated genome elimination based technology is developed to induce haploid production in plants. in this method, the function of centromere, is altered by manipulating the particular centromeric protein called cenh3 to produce haploid inducer lines. if these lines hybridized to the plant of the same species, the genome of the inducer line is eliminated and homozygous plants will achieve in one generation. we aim to isolate and characterize a kinetochore component protein “knl2” in solanum lycopersicum (slknl2). knl2, play a role in the cenh3 depositioning to the centromere in studied eukaryotes.methods: for cloning of slknl2, total rna of leaves were extracted and synthesized cdna was used as a template for pcr. subsequently, pcr products were cloned for sequencing using clonejet pcr cloning kit. in silico primary sequence analyses confirmed the successful cloning of slknl2.results: blast searches and alignment of slknl2 protein with other putative knl2 proteins revealed that nearly all of eukaryotes whose genomes are completely sequenced contain only a single copy of this gene, indicating an important function of it during evolution. slknl2 showed 37.4 percent identity to its orthologues protein in arabidopsis thaliana. in addition the two very important santa domain (play a role in regulating chromatin) and cenpc-like motif (essential for centromeric localization of knl2) were conserved in tomato as well. the editing of slknl2 gene will be done by crisper-cas9 technology to check whether disruption of its function could create inducer line for haploid production in tomato.conclusion: centromere-mediated genome elimination is a novel advancement in the production of haploid plant lines and manipulating the essential component of the kinetochore assembly could offer tremendous benefits in plant breeding programs.