Controls
We recommend using the following positive and negative controls for your miRNA modulation experiments.
Positive controls
miRNAs are expressed at various levels in different cell types; therefore, it can be difficult to find a good positive control. Ensure that the positive control that you select is expressed at sufficiently high levels to enable measurement of response.
An miRNA that can serve as a good positive control in many situations is miR-21-5p, which is conserved in many species and expressed at high levels in HeLa cells. Protein modulation can be confirmed by measuring
expression of endogenous miR-21 targets, such as PTEN [6] and PDCD4 [6–9], or using a reporter assay.
Species
| Mature miR-21-5p sequence* (copy and paste into IDT miRNA Inhibitor ordering tool) |
---|
Human, mouse, rat | uagcuuaucagacugauguuga
|
Negative controls
A good negative control should be inert and not modulate any genes in the system under study. This is difficult to achieve; however, we propose 2 negative control sequences that we have used throughout our product development and validation process, and
which we have found to work very well in vitro and in vivo.
Species
| Sequence* (copy and paste into IDT miRNA Inhibitor ordering tool) |
---|
NC1 Negative Control (human) | ucguuaaucggcuauaauacgc
|
NC5 Negative Control (human, mouse, rat) | accauauugcgcguauagucgc
|