shRNA vs siRNA
During the process of RNA interference (RNAi), expression of a target gene is knocked down with high specificity and selectivity. RNAi is a natural process, and it involves small interfering RNA (siRNA) and short hairpin RNA (shRNA) and bi-functional shRNA. Presently, RNAi is widely used as a tool for personalized cancer therapy. The applications of RNAi are basically done with the chemically synthesized double- stranded siRNA and vector based shRNA molecules. Although these two molecules have similar functional outcomes, they differ in their structure; thus, the molecular mechanisms of action, the RNA pathways, and off-target effects of these two molecules also differ.
shRNA is a sequence of small RNA molecule which makes a tight hairpin turn that can be used to silence a target gene expression during RNAi. Expression of shRNA is achieved by a vector, which can be either a virus or a bacterium or by delivery of plasmids. They are synthesized in the nucleus of cells and transported to the cytoplasm for further processes. These molecules have similar maturation pathways of miRNA; thus the synthesis of miRNA has provided the groundwork for understanding of shRNA synthesis. Either RNA polymerase II or III can transcribe shRNA through RNA polymerase II or III promoters. The advantage of the usage of shRNAs is that they have relatively low rate of degradation and turnover. The disadvantage is that it needs an expression vector, which may cause some safety issues.
siRNAs are double stranded RNA molecules composed of 20- 25 base pairs in length. These are used for gene suppression by silencing any gene with complementary nucleotide sequence in RNAi pathway. Gene knockdown by transfection of siRNA is often unsuccessful due the transient effect; especially in rapidly dividing cells and the suppression do not last longer. To overcome this issue, the siRNA is modified by introducing a short hairpin structure. This modified molecule then known as shRNA. shRNA should be converted to siRNA by a Dicer in order to proceed its normal function.
What is the difference between shRNA and siRNA?
• Unlike the siRNA, shRNA has an extra hairpin structure. shRNA is a modified version of siRNA.
• shRNA requires an expression vector, whereas siRNA does not.
• shRNA can be used for long term knockdown while siRNA can be used only for short term knockdown of genes.
• Unlike the gene suppression of siRNA, the suppression of shRNA is long lasting, and if it is inserted via an appropriate viral vector, it may produce permanent gene silencing effects.
• Dicer is required to convert the shRNA back to siRNA molecule in order to execute its normal functions.