Abstract
Targeted genome engineering (also known as genome editing) has emerged as an alternative to classical plant breeding and transgenic (GMO) methods to improve crop plants. Until recently, available tools for introducing site-specific double strand DNA breaks were restricted to zinc finger nucleases (ZFNs) and TAL effector nucleases (TALENs). However, these technologies have not been widely adopted by the plant research community due to complicated design and laborious assembly of specific DNA binding proteins for each target gene. Recently, an easier method has emerged based on the bacterial type II CRISPR (clustered regularly interspaced short palindromic repeats)/Cas (CRISPR-associated) immune system. The CRISPR/Cas system allows targeted cleavage of genomic DNA guided by a customizable small noncoding RNA, resulting in gene modifications by both non-homologous end joining (NHEJ) and homology-directed repair (HDR) mechanisms. In this review we summarize and discuss recent applications of the CRISPR/Cas technology in plants.
Highlights
Targeted genome engineering has emerged as an alternative to classical plant breeding and transgenic (GMO) methods to improve crop plants and ensure sustainable food production
Until recently the available methods have proven cumbersome. Both zinc finger nucleases (ZFNs) and TAL effector nucleases (TALENs) can be used to mutagenize genomes at specific loci, but these systems require two different DNA binding proteins flanking a sequence of interest, each with a C-terminal FokI nuclease module
A new method based on the bacterial CRISPR/Cas (CRISPR-associated) type II prokaryotic adaptive immune system [1] has emerged as an alternative method for genome engineering
Summary
Targeted genome engineering has emerged as an alternative to classical plant breeding and transgenic (GMO) methods to improve crop plants and ensure sustainable food production. Given that only a single RNA is required to generate target specificity, the CRISPR/Cas system promises to be more applicable to genome engineering than ZFNs and TALENs. Recently, eight reports describing the first applications of the Cas9/sgRNA system to plants have been published [9,10,11,12,13,14,15,16]. DNA targets and sgRNA guide sequences that differ from the canonical 20 bp length have been reported in some plant studies [10,12,13,14,15], while in the mammalian field targets of the consensus (N)20NGG are normally used.
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