Fig. 3

Gene insertion using CRISPR/Cas9 and plasmid-borne editing template in C. glutamicum SL4. a Schematic representation of pCas9, pgRNA4, and gene insertion event. b Colony counting and editing efficiency calculation of C. glutamicum SL4. pCas9 and pgRNA4 were co-transformed into C. glutamicum SL4 simultaneously and cells were spread on SGY plates supplemented with Km, Cm, and IPTG (1 mM) immediately after recovery. Colonies on the selective plates were counted and verified using colony PCR. c PCR verification of rfp insertion in C. glutamicum SL4 using the primer pair (rfp-up and ldhA-down) shown in a. M, DNA marker; –, wild-type control; 1–10, ten small colonies. This displayed the result of Replicate 1 in b. The results of Replicate 2 and Replicate 3 were shown in Additional file 1: Figure S4