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Fig. 4 | Biotechnology for Biofuels

Fig. 4

From: Endogenous CRISPR-assisted microhomology-mediated end joining enables rapid genome editing in Zymomonas mobilis

Fig. 4

Isolation of a sigma factor encoding gene deletion mutant via CRISPR interference and MMEJ repair. a Colony PCR screening for MMEJ mutants transformed with the self-interference plasmid against the ZMO0626 using primers located upstream or downstream of the target site. Predicted sizes of PCR products from the wild type (wt) and the MMEJ-mediated deletion mutants (del) are indicated, respectively. M, DNA size marker. b MMEJ-mediated deletions with different microhomology repeats adjacent to the deletion regions at the σ28 factor encoding gene ZMO0626. A blue bar on the gene indicates the protospacer location. Grey bars indicate the sequenced regions, white bars and the numbers at the right indicate deletion regions and deletion length (bp). c Colony PCR screening for the homozygous deletion mutant colonies isolated by a one-round spreading of the transformant corresponded to lane 1 indicated by a red arrow in a. d Agar streak test of the mobility of the wild type and the ZMO0626 mutant. The ZMO0626 gene encodes a sigma factor (σ28) that regulates the transcription of flagella-related genes

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