Presence of selected tomato (Solanum lycopersicon) microRNAs (sly-miRNAs) was validated and their expression profiles established in roots, stems, leaves, flowers and fruits of tomato variety Jiangshu14 by quantitative RT-PCR (qRT-PCR). In addition conservation characteristics these sly-miRNAs were analyzed and target genes predicted bioinformatically. Results indicate that some of these miRNAs are specific to tomato while most are conserved in other plant species. Predicted sly-miRNA targets genes were shown to be targeted by either by a single or more miRNAs and are involved in diverse processes in tomato plant growth and development. All the 36 miRNAs were present in the cDNA of mixed tissues and qRT-PCR revealed that some of these sly-miRNAs are ubiquitous in tomato while others have tissue-specific expression. The experimental validation and expression profiling as well target gene prediction of these miRNAs in tomato as done in this study can add to the knowledge on the important roles played by these sly-miRNAs in the growth and development, environmental stress tolerance as well as pest and disease resistance in tomatoes and related species. In addition these findings broaden the knowledge of small RNA-mediated regulation in S. lycopersicon. It is recommended that experimental validation of the target genes be done so as to give a much more comprehensive information package on these miRNAs in tomato and specifically in the selected variety.