Scientists Develop DNA- and Marker-free Genome-editing System using Zygotes in Rice

The introduction of CRISPR-Cas9 system has led to the development of rapid and cost-effective procedures to generate new mutant populations in plants. Even if genome-edited plants from several species have been developed successfully using a technique wherein a Cas9-guide RNA (gRNA) expression cassette and selectable marker are integrated into the genomic DNA through Agrobacterium tumefaciens-mediated transformation or particle bombardment, CRISPR-Cas9 system increases the chance for off-target modifications, and foreign DNA sequences cause legislative concerns about GMOs. Thus, DNA-free genome editing has been developed, which involves the transport of preassembled Cas9-gRNA ribonucleoproteins (RNPs) into protoplasts derived from somatic tissues by polyethylene glycol-calcium mediated transfection in tobacco, Arabidopsis, rice, potato, etc. or into embryo cells thru the use of a gene gun in maize and wheat.

Experts from RIKEN Cluster for Science, Technology and Innovation Hub reported in Nature Plants a genome-editing system via direct delivery of Cas9–gRNA RNPs into plant zygotes. Cas9–gRNA RNPs were integrated into rice zygotes produced by in vitro fertilization of isolated gametes and the zygotes were cultured into mature plants in the absence of selection agents, resulting in the regeneration of rice plants with targeted mutations in around 14 to 64 percent of plants.

Based on the study, the new plant genome-editing system has a huge potential for the continuous development of rice and other vital crops.

Read the abstract in Nature Plants.