Figure 6

Mechanism of deletion of the main folate transporter gene FT1 in L. major selected for MTX resistance. (a) A portion of the L. major chromosome 10 showing some of the FT genes. Approximate location of PvuI sites (crosses) and their size are shown. Primers F and R are indicated by half arrows. The relative hybridization data obtained from RNA expression profiling (RNA) and comparative genomic hybridization (DNA) are shown. Due to conservation between the FT genes, the 70-mer probes for LmjF10.0380, FT1 and LmjF10.0390 are not discriminatory. (b) Southern blot of Leishmania total DNA digested with PvuI and hybridized to a probe recognizing conserved sequences of most FBT genes (indicated by bars underneath the genes in (a,c)). The genes corresponding to some hybridizing bands are indicated. (c) Model for the deletion of FT1 mediated by the homologous recombination of the conserved sequences between the folate transporter genes LmjF10.0380 and LmjF10.0390 (Figure S3 in Additional data file 2). (d) PCR with primers F and R to support the model shown in (c). Lane 1, L. major wild-type cells; lane 2, L. major MTX60.4.