Extended Data Fig. 2: Evaluation of the efficiency and precision of the tRNA-mediated gRNA processing by cRT-PCR.
From: Highly efficient DNA-free plant genome editing using virally delivered CRISPR–Cas9
a, Schematic representation of gRNA circularization and terminal sequence mapping. The divergent arrow pairs denote annealing positions of abutting primers used for cRT-PCR. The hypothesized size of each amplified fragment are indicated. A(n), poly(A) tails with undefined base number. b, cRT-PCR analysis of the gRNA processing efficiency. Arrow denotes DNA fragments corresponding in size to mature gRNA, and asterisk labels the unprocessed 5′ UTR-gRNA-3′ UTR fragment. M, 500-bp ladder marker. c, Sanger sequencing of the gRNA-like cRT-PCR products. The linear sequences are aligned with a canonical gRNA (WT). The sequences of gRNA spacer, scaffold, tRNA 5′ leader, and SYNV UTR are shown in blue, black, purple, and red color letters, respectively. x# shown on the right indicate the number of individual colonies with the identical sequence.
