Figure 2 | Scientific Reports

Figure 2

From: Establishment of a modified CRISPR/Cas9 system with increased mutagenesis frequency using the translational enhancer dMac3 and multiple guide RNAs in potato

Figure 2

Construction of the CRISPR/Cas9 vector. (A) Structure of the potato GBSSI gene and the target site of guide RNAs. Boxes indicate the region for exons. Arrows indicate the locations of the guide RNAs. (B) Construction of the multiple gRNA genes. This figure shows a scheme of construction on the case using two gRNAs, gRNA #1 and #3 as the representative guide RNAs. DNA fragments corresponding to the target sequences were chemically synthesized and each of them was inserted between the AtU6 promoter and gRNA scaffold in a pMR vector. The resultant gRNA genes were introduced into pBS_GwIsceI using the multi-Gateway method. AtU6-26: Arabidopsis thaliana U6-26 promoter; gRNA scaffold: region for the gRNA scaffold; and attL1 and attL5 and attR1 and attR2: representative att regions for Gateway recombination sites. (C) Construction of the CRISPR/Cas9 vector plasmids. This figure shows the CRISPR/Cas9 vector plasmid, which contains an appropriate gRNA gene and the Cas9 gene with dMac3. gRNAs: representative gRNA genes. Other elements are cited from those in Fig. 1.

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