Targeted genome editing : an innovative system for rice and other crop improvement

Escalating human population as well as in extreme weather events require the sustaining improvement of crop varieties especially in rice. A direct, rapid, and low-cost method is needed for supporting high-yield, multi-stress resistant rice varieties. Conventional rice breeding and genetic modification (GM) methods have advantages in capturing favorable agricultural traits but takes a very long time. However, the targeted genome editing technology of key functional genes promises to be a powerful tool in accelerating rice varietal improvement. Targeted genome engineering (also known as genome editing) has emerged as an alternative to classical plant breeding and GM technology methods to improve rice crop. In the past years, common available tools for introducing site-specific double strand DNA breaks were zinc finger nucleases (ZFNs) and TAL effector nucleases (TALENS). However, these technologies have not been widely adopted by the 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)/Cas9 (CRISPR-associated) immune system. The CRISPR/Cas9 system allows targeted cleavage of genomic DNA guided by a customizable small non-coding RNA, resulting in gene modifications by both non-homologous and joining (NHEJ) and homology-directed repair (HDR) mechanisms. And by mendelian genetics, rice events with targeted mutation can be considered as a non-GM rice crop when there is no more traces of t-DNA resulted by segregation on the next generation. In the present study, the authors transformed IR64 mega variety by Agrobacterium-mediated gene transfer with CRISPR/Cas9 construct pRGEB31 (Xie et al. 2013) targeting the elF4G locus region (LOC_Os 07G36940). Based on the number of transgenic rice produced, the authors are currently analyzing the efficiency of the CRISPR/Cas9 system in targeting the elF4G locus region through T7 endonuclease assay.