Figure 3 | Scientific Reports

Figure 3

From: CRISPR/Cas9 mediated targeting of multiple genes in Dictyostelium

Figure 3

Targeting of two developmental genes using CRISPR. (A) Schematic overview of the all-in-one CRISPR/Cas9 vector, pTM1285. A pair of annealed oligonucleotides is assembled into the all-in-one vector using the Golden Gate assembly method. Amp, ampicillin resistance gene; act15, act15 promoter; act8, act8 terminator; tRNA, isoleucine tRNA, mhcA, mhcA terminator; neo, neomycin resistance gene; act6, act6 promoter. (B) Schematic view of sgRNA targeting locus of the acaA and pkaC genes. The blue sequences indicate the target sites of the sgRNA. (C) Frequency of aggregation-negative and mutated clones. (D) Wild-type sequence for acaA and pkaC and sequences derived from 4 independent positive clones by mutation detective PCR are presented. The target sequence is shown in blue, and mutations are shown in red. Numbers in parentheses indicate the number of modified nucleotides.