172. PROGRAMMABLE RNA RECOGNITION WITH CRISPR/CAS9

Department: Mechanical & Aerospace Engineering
Research Institute Affiliation: Graduate Program in Materials Science and Engineering
Faculty Advisor(s): Gene Yeo

Primary Student
Name: David Allen Nelles
Email: dnelles@ucsd.edu
Phone: 858-534-9322
Grad Year: 2016

Abstract
RNA-programmed genome editing using CRISPR/Cas9 from Streptococcus pyogenes (spCas9) has enabled rapid and accessible alteration of any genomic locus in a variety of organisms. As a result, a genome engineering revolution supported by spCas9 is expected to provide unparalleled means to correct genetic disease and design custom organisms. In contrast, there is currently no accessible means to target and alter arbitrary RNAs in living cells. An analogous means to target RNA would allow alteration and imaging of endogenous RNA transcripts in order to assess or alter RNA processing in healthy and diseased cells. But there currently exist no flexible means to target an RNA-of-choice. Recent work has demonstrated the ability of spCas9 to bind RNA in purified cell extracts while avoiding encoding DNA and we have recently achieved recognition of RNA in living cells programmed by a pair of nucleic acids in combination with nuclease-inactive spCas9 fused to fluorescent proteins. This approach supports tracking of endogenous RNAs with RNA-targeted Cas9 (RCas9) signal distributions that correlate highly with fluorescence in situ hybridization. We have also tracked RNA translocation in living cells to stress granules. Our results establish RCas9 as a means to bind and track RNA in living cells in a programmable manner without the requirement of genetically encoded tags and indicates the potential of spCas9 as a means to manipulate the transcriptome in addition to the genome.

Industry Application Area(s)
Life Sciences/Medical Devices & Instruments

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