Skip to main content
Fig. 3 | Parasites & Vectors

Fig. 3

From: LmABCB3, an atypical mitochondrial ABC transporter essential for Leishmania major virulence, acts in heme and cytosolic iron/sulfur clusters biogenesis

Fig. 3

Generation and characterization of L. major promastigotes with one LmABCB3 allele deleted. a Schematic representation of the LmABCB3 locus and the hygromycin-resistance gene targeting construct used for gene replacement. The primers used (arrows 1–3) to verify the specific gene targeting and the expected sizes of the PCR-amplified products with the different pairs of primers are indicated. b PCR analysis of the LmABCB3 locus in control and single knock out (LmABCB3 +/−) mutant promastigotes. The specific gene-targeting PCR product (primers 1 and 3, 3.6 kb) confirmed that replacement with the hygromycin-resistance gene in one LmABCB3 allele in LmABCB3 +/−parasites (+/−, lanes 2). Primers 1 and 2 amplified the expected 1.6 kb product in Wt (+/+, lanes 1) and in LmABCB3 +/−parasites. Lanes 3 shows the DNA marker used. c LmABCB3 +/−promastigotes have reduced LmABCB3 expression. The expression level of LmABCB3 from control and LmABCB3 +/− L. major promastigotes was analyzed by qRT-PCR using mRNA isolated from each cell line as described in Methods. **p <0.001. d LmABCB3 +/− parasites grow as axenic promastigotes. Growth curve obtained after cultivation of control (white circles) and LmABCB3 +/−(black circles) promastigotes during the indicated time. The results represent the mean ± SEM of three independent experiments. *p < 0.05; **p < 0.001

Back to article page