In vivo interrogation of gene function in the mammalian brain using CRISPR-Cas9
By L. Swiech, Matthias Heidenreich, Abhishek B. Banerjee, Naomi Habib, Yinqing Li, John J. Trombetta, Mriganka Sur, Feng Zhang. Nature Biotechnology, 2014
764 citations recorded by the source corpus.
Probing gene function in the mammalian brain can be greatly assisted with methods to manipulate the genome of neurons in vivo. The clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated endonuclease (Cas)9 from Streptococcus pyogenes (SpCas9) can be used to edit single or multiple genes in replicating eukaryotic cells, resulting in frame-shifting insertion/deletion (indel) mutations and subsequent protein depletion. Here, we delivered SpCas9 and guide RNAs using adeno-associated viral (AAV) vectors to target single (Mecp2) as well as multiple genes (Dnmt1, Dnmt3a and Dnmt3b) in the adult mouse brain in vivo. We characterized the effects of genome modifications in postmitotic neurons using biochemical, genetic, electrophysiological and behavioral readouts. Our results demonstrate that AAV-mediated SpCas9 genome editing can enable reverse genetic studies of gene function in the brain.
Subfields: Molecular Biology
View Paper: https://www.semanticscholar.org/paper/71c32f87b7f51d76ee9b8161809683debe1ecad3
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Citations in the corpus
- Transcriptome engineering with RNA-targeting Type VI-D CRISPR effectors (2018, neutral): “7 kb) makes it challenging to effectively deliver many single effector CRISPR enzymes (Abudayyeh et al., 2017; Ran et al., 2015; Swiech et al., 2015).”
- Multiplex gene editing by CRISPR-Cpf1 through autonomous processing of a single crRNA array (2016, methodology): “This plasmid also included an green fluorescent protein (GFP) fused to the KASH nuclear transmembrane domain 4 to enable FACS of targeted cell nuclei 5 .”
- Delivering CRISPR: a review of the challenges and approaches (2018, background): “Many groups have reported success packaging the SpCas9 and sgRNA into two separate AAV particles and using them for co-infection (Swiech et al., 2015; Hung et al., 2016).”