RNA-programmed genome editing in human cells

Abstract

Type II CRISPR immune systems in bacteria use a dual RNA-guided DNA endonuclease, Cas9, to cleave foreign DNA at specific sites. We show here that Cas9 assembles with hybrid guide RNAs in human cells and can induce the formation of double-strand DNA breaks (DSBs) at a site complementary to the guide RNA sequence in genomic DNA. This cleavage activity requires both Cas9 and the complementary binding of the guide RNA. Experiments using extracts from transfected cells show that RNA expression and/or assembly into Cas9 is the limiting factor for Cas9-mediated DNA cleavage. In addition, we find that extension of the RNA sequence at the 3′ end enhances DNA targeting activity in vivo. These results show that RNA-programmed genome editing is a facile strategy for introducing site-specific genetic changes in human cells.DOI: http://dx.doi.org/10.7554/eLife.00471.001

Oligonucleotide-based targeted gene editing in C. elegans via the CRISPR/Cas9 system

“…Compared with the zinc finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN), which have been used for genome editing [1], the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/ CRISPR-associated (Cas) system has emerged as a new powerful tool for genome modifications. It has recently been adopted for genome editing in human cell lines [2][3][4], mouse [5], zebrafish [6], C. elegans [7][8][9][10][11][12], and plants [13].…”

Genome Editing in Neuroepithelial Stem Cells to Generate Human Neurons with High Adenosine-Releasing Capacity

“…However, translation of these findings to human cells was, for a long time, complicated by the fact that homologous recombination in human cells proved to be highly inefficient 35. With the recent advent of genome editing technologies such as ZFNs 36, transcription activator‐like effector nucleases 37, and CRISPR‐Cas/RNA‐guided nucleases 38, gene editing has become broadly applicable to human cells. Typically, these techniques are applied to human pluripotent stem cells, which are then used to derive defined somatic cell types required for the individual application.…”

CRISPR, the disruptor

“…CRISPR is used in mouse and human cells, fuelling rapid uptake of the technique by researchers [11][12][13] .…”