From wall hung TVs to the latest wearable; displays have been a key technology differentiator for years. Now we’d like to introduce you to CRISPR-Display, one of the latest applications of CRISPR technology for the lncRNA lover that wants to elevate their functional game.
Interest in lncRNAs continues to surge, but like other uncharted areas of research, loads of functional studies are still needed. Now, by utilizing dCas9 and some designer sgRNA, researchers have access to a lncRNA delivery system with more localization power than the last 4 digits on your zip code.
CRISPR-Display comes to you from the talented team of lncRNA researchers in the Rinn lab at Harvard.
Here are the specs on CRISPR-Display:
- The sgRNA is upgraded with custom RNA domains (outside targeting sequence) to create the substrate needed for 3 possible distinct functional domains that protrude from the CRISPR/sgRNA complex
- These domains are located 5’, center, and 3’.
- dCas9 and sgRNAs containing the modifications are delivered via lentiviral vector to mammalian cells (HEK293FT cells).
- The system was tested using a VP64 activation domain to drive luciferase expression.
- The results were also confirmed using RNA Immunoprecipiation (RIP)
- The custom RNA cargo put into the domains can form functional secondary structure to be ‘displayed’ and can be up to 4.8 kb.
- The functionality of the loop domains was demonstrated in displays that included natural lncRNAs, protein-binding cassettes that allow for bridged activation, artificial aptamers, and pools of random sequences.
- Multiple display functions can be carried out at different loci in a single cell.
CRISPR-Display enables new experimentation by allowing for the precision targeting of functional RNA domains and ribonucleoproteins to loci of interest. Ultimately, CRISPR-Display enables not only the study of natural RNA domains but also synthetic domains for custom RNA devices, which effectively brings ncRNA into the CRISPR renaissance.
Go see the whole display at Nature Methods, June 2015