Figure 1. The CRISPR/spCas9 strategy for mutating binding motifs for transcription factors.
(A) The human β-globin locus is presented. Vertical gray arrows indicate DNase I HSs in the LCR and 3′-HS1. The five globin genes, ε, Gγ,Aγ, δ, and β are represented by rectangle in sequence. DNA sequences are for the LCR HS3 and HS2, and the binding motifs of transcription factors are marked by colored bases; GATA-1 motif: red, KLF1 motif: green, and TAL1 motif: blue. Target sequences for sgRNA are indicated by purple lines with PAMs in red lines. (B) Guide sequences of sgRNA are listed with names, scores, and PAM sequences, where (+) and (–) mean guide sequences are positive or negative strand DNA, respectively. (C) Experimental procedure for mutating binding motifs for transcription factors in MEL/ch11 cells is illustrated and described in ‘Materials and methods’ section.