Figure 4.

A novel strategy for highly efficient CRISPR/Cas9 targeted genomic deletions in Arabidopsis. (A) The Cas9 expression vector was integrated into indRKD4 plants using floral dipping. Positive transformants were selected using the fluorescent seed-specific reporter. Positive seeds were grown in liquid medium containing dexamethasome. After two weeks cell suspensions were transferred to plates containing standard medium. Regenerated plants were tested for Cas9 nuclease activity by PCR and selection of heritable deletion was determined in plants lacking Cas9 activity. Primers JD157 with JD158 and JD125 with JD126 were used for the ISU1 deletion and At5g60390 control, respectively. (B) PCR analysis of the ISU1 locus to identify targeted deletions. (C) Sanger sequencing results of plants containing independent ISU1 heritable deletions.