Invention:
MicroRNAs (miRNAs) are regulatory RNA molecules that bind to target messenger RNAs (mRNAs) and affect the stability or translational efficiency of the bound mRNAs. Single or dual-luciferase reporter systems have been successfully used to identify miRNA target genes in mammalian cells, however, they are not sensitive enough to verify miRNA–target gene relationships in insect cell lines. The promoters for the target luciferase in insect cells are too weak to drive sufficient expression of the target luciferase in insect cells.
This technology replaces the promoter used for the mammalian cells with a different promoter that has the potential to bind in insect cells. The promoter psiCHECK-2-TK has suitable insect cell luciferase activity ratios regardless of the cell line. PsiCHECK-2-TK was able to detect the interaction between Helicoverpa armigera miRNA9a and its target in both mammalian and H. zea cell lines; furthermore, psiCHECK-2-TK with the target sequence, HzMasc accurately differentiated between H. zea cell lines with or without the negative regulation factor of HzMasc. This demonstrates that psiCHECK-2-TK can be used to functionally characterize small RNA target genes in both mammalian and insect cells.
Link to published paper: https://doi.org/10.1111/1744-7917.12945
Background:
Small non-coding RNAs, such as microRNAs (miRNAs), are important for regulating gene expression. miRNAs regulate many cellular processes including proliferation, differentiation, reproduction, death, metabolism, and development. miRNAs will interact with its target gene through base pairing, but it is very specific and there needs to be a minimum of eight similar base pairs. Currently, there is only one commercially available dual-luciferase (a photoprotein that is bioluminescent when bonded to its target) reporter system that can verify RNA genes. With only one dual-luciferase on the market there is a possibility of many miRNAs that are not being identified do the specificity required. Having the technology to alter and bind to these miRNAs can be useful for the pesticide industry for the development of a new pesticide.
Applications:
- Insect cell line research
- Pesticides
Advantages:
- Can be commercialized
- Can be repeated to find more sequences
