Hi Kartik,

miRNA are biological, I mean that that's the way cells are regulating gene expression in specific conditions, while shRNA (or siRNA, crispr-cas9...) are artifactual and targeted gene silencing. Where one miRNA can have lot of targets, one shRNA should have only one, and the quantity of this tool can be controlled.

fred

Hi Kartik,

The gene-silencing from RNAi molecules (e.g., miRNA, siRNA, shRNA) is due to a single-strand mature RNA (guide strand) guiding the RISC (RNA induced silencing complex). The mature strand of all RNAi molecules are derived from a duplex RNA.

As Frederic mentioned miRNA are natural molecules produced in the body. In nature, the miRNA duplexes are produced from a stem-loop structure called the precursor-miRNA.

Here is a nice review article on miRNA biogenesis: Finnegan, E.F. and Pasquinelli, A.E., 2013, DOI: 10.3109/10409238.2012.738643.Article MicroRNA biogenesis: Regulating the Regulators

To study transient effects on gene-silencing, miRNA-mimics or siRNAs are used.

- The miRNA-mimics have the same sequence as natural miRNAs.

- The sequence of siRNAs (21 base-pairs) are carefully designed to be fully complementary to a target gene and minimize binding to other genes.

shRNAs (short-hairpin RNAs) resemble the stem-loop structure of precursor-miRNA. Either miRNA or siRNA sequence is introduced in the stem-region. Initially, miRNA and siRNAs were synthesized as shRNAs. Currently, most of the commercially available miRNA-mimics and siRNAs are available as a duplex RNAs.

The advantage of shRNAs is that it undergoes DICER processing, which can increase RISC loading, as reported for a synthetic DICER-substrate 27mer siRNA (Rose et al., 2005, DOI: 10.1093/nar/gki732 Article Functional polarity is introduced by Dicer processing of sho...

Limitation of shRNAs: DICER and RISC are limited cellular resources. It is critical for the normal functioning of the cell. Hence, at higher concentrations, shRNA saturates DICER and RISC leading to toxicity (Grimm, D., 2011, Silence 2, 8, DOI: 10.1186/1758-907X-2-8; Article The dose can make the poison: Lessons learned from adverse i...

- This limitation is also true for siRNAs and miRNAs, but under in-vitro conditions, the concentration can be tailored to have minimal toxicity and maximal gene-silencing.

Difference between miRNA and siRNA:

In nature, miRNA-mediated mRNA-recognition happens mainly due to the base-pairing between 6 - 8mers (seed-sequence) between the miRNA and the target mRNAs. Due to the short recognition length requirement, miRNA binds to multiple mRNAs, hence down-regulates multiple genes. This also results in decreased efficiency of gene-silencing for any given gene compared to siRNA.

- In contrast, as siRNAs are designed specifically to bind to a single gene (21-base-pairing), the silencing efficiency is usually higher for that single gene. However, due to the 6-8mer miRNA-like base-pairing, siRNAs can down-regulate multiple genes too at excessive concentrations, which is referred as off-target effects.

To study long-term effects, shRNAs are encoded into a DNA and incorporated into the genome under the control of constitutive or variable promoter of choice.

Best wishes.

siRNA or shRNA will induce cleavage of the target mRNA within a perfect duplex (so in a perfect world- just one target, and the outcome is knockdown- mRNA and protein levels go down by eg 90%); microRNA binds many mRNAs with a number of mismatches, and in many cases suppresses protein production (just by a small percentage eg 10%) rather than induce cleavage of the target.