Repurposing of Anthocyanin Biosynthesis for Plant Transformation and Genome Editing

Abstract

CRISPR/Cas9 gene editing technology has been very effective in editing genes in many plant species including rice. Here we further improve the current CRISPR/Cas9 gene editing technology in both efficiency and time needed for isolation of transgene-free and target gene-edited plants. We coupled the CRISPR/Cas9 cassette with a unit that activates anthocyanin biosynthesis, providing a visible marker for detecting the presence of transgenes. The anthocyanin-marker assisted CRISPR (AAC) technology enables us to identify transgenic events even at calli stage, to select transformants with elevated <i>Cas9</i> expression, and to identify transgene-free plants in the field. We used the AAC technology to edit <i>LAZY1</i> and <i>G1</i> and successfully generated many transgene-free and target gene-edited plants at T1 generation. The AAC technology greatly reduced the labor, time, and costs needed for editing target genes in rice.