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Chemically modified guide RNAs enhance CRISPR-Cas13 knockdown in human cells
Cell Chemical Biology ( IF 8.6 ) Pub Date : 2021-08-02 , DOI: 10.1016/j.chembiol.2021.07.011 Alejandro Méndez-Mancilla 1 , Hans-Hermann Wessels 1 , Mateusz Legut 1 , Anastasia Kadina 2 , Megumu Mabuchi 3 , John Walker 2 , G Brett Robb 3 , Kevin Holden 2 , Neville E Sanjana 1
更新日期：2021-08-02
Cell Chemical Biology ( IF 8.6 ) Pub Date : 2021-08-02 , DOI: 10.1016/j.chembiol.2021.07.011 Alejandro Méndez-Mancilla 1 , Hans-Hermann Wessels 1 , Mateusz Legut 1 , Anastasia Kadina 2 , Megumu Mabuchi 3 , John Walker 2 , G Brett Robb 3 , Kevin Holden 2 , Neville E Sanjana 1
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RNA-targeting CRISPR-Cas13 proteins have recently emerged as a powerful platform to modulate gene expression outcomes. However, protein and CRISPR RNA (crRNA) delivery in human cells can be challenging with rapid crRNA degradation yielding transient knockdown. Here we compare several chemical RNA modifications at different positions to identify synthetic crRNAs that improve RNA targeting efficiency and half-life in human cells. We show that co-delivery of modified crRNAs and recombinant Cas13 enzyme in ribonucleoprotein (RNP) complexes can alter gene expression in primary CD4+ and CD8+ T cells. This system represents a robust and efficient method to modulate transcripts without genetic manipulation.
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