Aims: To address the possible use of RNA interference in diagnosis, prognosis and therapy of various diseases and explain the obstacles in RNA interference based therapy. Methods: Review of the literature. Results and discussion: The MicroRNAs (miRNAs) were shown to play a role in various normal and pathological biological processes, i.e. the regulation of cell cycle and apoptosis, cell differentiation, wound healing process, immune system, etc. Furthermore, abnormal expression of miRNA was found in various diseases. Therefore, when the sample can be easily collected, miRNA expression profile can be used to detect diseases where early diagnosis is advantageous, such as in various malignancies or diseases that show myriads of symptoms such as autoimmune diseases. Further, different expression of miRNA in tumour subtypes can be used to predict the subtypes and hence the prognosis. For therapy, small (short) interfering RNAs (siRNAs) can be developed to 'switch off' up-regulated genes or miRNAs or the suppressors of down-regulated genes or miRNAs. These approaches in various animal models of diseases showed promising results and human trials for viral infections are underway. However, obstacles to the application in human might be encountered, such as degradation of the siRNAs before it can exert its function, target cell penetration and 'off target' toxic effects. Still, it is believed that modification of the RNA, development of carrier vehicles and mode and route of administration might solve these obstacles. Conclusion: MicroRNAs profile differences between normal and pathological condition might be promising as biomarker in early diagnosis and prognosis, while siRNA showed promising result in the therapy of various diseases in animal models.