CRISPR-Based Technologies for the Manipulation of Eukaryotic Genomes

Abstract

The CRISPR-Cas9 RNA-guided DNA endonuclease has contributed to an explosion of advances in the life sciences that have grown from the ability to edit genomes within living cells. In this review we summarize CRISPR-based technologies that enable mammalian genome editing and their various applications. We describe recent developments that extend the generality, DNA specificity, product selectivity, and fundamental capabilities of natural CRISPR systems, and some of the remarkable advancements in basic research, biotechnology, and therapeutics development that these developments have facilitated.

Human Germline Genome Editing

“…CRISPR/Cas9 and other genome-editing methods have been thoroughly reviewed elsewhere. [1][2][3] Like many other robust DNA modification technologies, CRISPR/Cas9 has quickly become a widely used research tool, and its embrace testifies to the ease with which it can be customized and its effectiveness in multiple cell types and species. In many ways, preceding gene-transfer technologies that fell short of ''genome editing''-i.e., introduced genes into cells but did not permanently incorporate them into the genome-laid the groundwork for the issues presented in this statement.…”

Future Perspectives for Cancer Therapy Using the CRISPR Genome Editing Technology

“…The elucidation of the CRISPR-cas-based gene editing machinery and understanding its mode of operation, led to the development of a CRISPR-cas-based biotechnological tool which enables the manipulation of eukaryotic genomes [18][19][20][21] and is useful in numerous applications in basic science, biotechnology and medicine [22]. Among its many potential applications, the CRISPR-cas9 technology can help correct genetic defects, prevent the spread of a wide range of diseases, and restore and enhance the immune system's ability to fight cancer.…”

Versatile and robust genome editing with Streptococcus thermophilus CRISPR1-Cas9

“…Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins form a prokaryotic adaptive immune system and some of its components have been harnessed for robust genome editing 1 . Type II-based editing tools rely on a large multidomain endonuclease, Cas9, guided to its DNA target by an engineered single-guide RNA (sgRNA) chimera 2 (See 3,4 for a classification of CRISPR-Cas systems).…”

“…While Streptococcus pyogenes (SpCas9) remains the most widely used Cas9 variant for genome engineering, the diversity of naturally occurring RNA-guided nucleases is astonishing 4 . Hence, Cas9 enzymes from different microbial species can contribute to the expansion of the CRISPR toolset by increasing targeting density, improving activity and specificity as well as easing delivery 1,6 .In principle, engineering complementary CRISPRCas systems from distinct bacterial species should be relatively straightforward, as they have been minimized to only two components. However, many such enzymes were found inactive in human cells despite being accurately reprogrammed for DNA binding and cleavage in vitro [7][8][9][10] .…”