Fig. 2: CRISPR/Cas9-mediated mutations were identified with high-throughput sequencing analysis of the PCR-amplified sgRNA-target region of the pds-12 mutant. | Horticulture Research

Fig. 2: CRISPR/Cas9-mediated mutations were identified with high-throughput sequencing analysis of the PCR-amplified sgRNA-target region of the pds-12 mutant.

From: A method for the production and expedient screening of CRISPR/Cas9-mediated non-transgenic mutant plants

Fig. 2

The red-marked TGG sequence on the x axis represents the three PAM nucleotides. Black circles represent nucleotide variant frequencies (NVF) of PCR product amplified from wild-type (WT) DNA. Red triangles represent the NVF of PCR product amplified from the genomic DNA of a 42-plant pool containing the pds-12 mutant line, displaying much higher NVF at positions 45–51. Blue squares represent the NVF of the same PCR products as the red triangles, but diluted six times with the WT PCR product, demonstrating that the NVF at nucleotide positions 45–51 are significantly reduced following the dilution. Using the WT PCR product dilution method, mutations at positions 45–51 were identified, consistent with the DNA sequencing results of the pds-12 mutant plant (Table 2)

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