Figure 1 : Characterization of MtbΔkefB mutant.

From: KefB inhibits phagosomal acidification but its role is unrelated to M. tuberculosis survival in host

Figure 1

(A) Representation of the genomic arrangement of the disrupted kefB gene and the depiction of the primer pairs employed for the characterization. (B) Confirmation of kefB gene deletion in M. tuberculosis by PCR by using the gene specific primers KefB-F-NdeI and KefB-R-NdeI (marked by red arrows in Fig. 1A) to obtain 1.1 kb amplification in M. tuberculosis (lane 3) and 2.2 kb amplification in MtbΔkefB (lane 4). 100 bp ladder and λHindIII were loaded in lanes 1 and 2, respectively. (C) Confirmation of kefB gene deletion in M. tuberculosis by PCR by using the primers KefB-Dn and Hyg-Dn (marked by green arrows in Fig. 1A). Lane 1 shows a 909 bp amplification in MtbΔkefB, lane 2 – 100 bp ladder (D) Confirmation of kefB gene deletion in M. tuberculosis by PCR by using the primers KefB-up and Hyg-up (marked by blue arrows in Fig. 1A). Lane 1 – 100 bp ladder and lane 2 shows a 961 bp amplification in MtbΔkefB. (E) Confirmation of complementation of the M. tuberculosis kefB mutant. The presence of pVR1-prokefB was confirmed by restriction digestion of the plasmid isolated from the complemented strain to yield a 1.1 kb band of kefB gene along with its promoter (lane 3). λHindIII and 100 bp ladder were loaded in lanes 1 and 2, respectively.