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. 2018 Sep 25;8:14355. doi: 10.1038/s41598-018-32702-w

Figure 3.

Figure 3

Marker-less RNP-CRISPR-Cas9 gene targeting of ALB1 without donor DNA. (a) Transformation plates from ALB1-targeting RNP-CRISPR-Cas9 without donor DNA showing rare albino patches (b) Primary colonies obtained by picking from the white patches shown in a. (c) Purified regenerants following a combination of subculture (hyphal tip isolation) and single spore isolation in most cases gave pure albino colonies. (d) Gel electrophoresis of the PCR products generated using the genomic DNA of the purified albino regenerants and primers PKS-ck-F and PKS-ck-R which flank the ALB1-targeting RNP-CRISPR-Cas9 genomic target sequence showing visible variation in product size. The gel image shown here was cropped, but no other bands were present. The full gel is given in Fig. S1. (e) Sequences of the amplicons shown in (d), showing a range of mutations and indels.