Several expression systems for multiple gRNAs have been developed in the
CRISPR-Cas9 system to induce multiple-gene modifications in plants. The team
of Ryosuke Hashimoto from Tokushima University in Japan evaluated the
mutation efficiencies in the tomato genome using multiplex CRISPR-Cas9
vectors consisting of various Cas9 expression promoters with multiple gRNA
expression combinations.

In tomato calli induced with these vectors, mutation patterns varied
depending on the promoters used to express Cas9. Notably, a high efficiency
of mutations was achieved in the tomato genome when the Cas9 was driven by
the tomato ELONGATION FACTOR-1á (SlEF1á) promoter.

These results suggest that optimizing the Cas9 expression promoter used in
CRISPR-Cas9 can improve multiplex genome editing.

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