Jr. Webb et Wr. Mcmaster, LEISHMANIA-MAJOR HEXBP DELETION MUTANTS GENERATED BY DOUBLE TARGETED GENE REPLACEMENT, Molecular and biochemical parasitology, 63(2), 1994, pp. 231-242
The Leishmania major single-stranded DNA binding protein HEXBP contain
s nine 'CCHC' zinc finger motifs and binds to oligodeoxynucleotides de
rived from the antisense strand of the GP63 gene 5' flanking region in
gel mobility shift assays and UV-crosslinking assays. In the present
study a HEXBP-deficient clone of L. major was generated by double targ
eted gene replacement. The two HEXBP alleles were sequentially replace
d with genes encoding resistance to the aminoglycoside antibiotics hyg
romycin B and G418 and drug-resistant clones were selected by plating
on semi-solid drug-containing media. Successful deletion of both copie
s of the HEXBP gene implies that HEXBP is a not essential for growth o
f Leishmania promastigotes. Characterization HEXBP-deficient promastig
otes revealed that HEXBP deficiency had no effect on the abundance of
GP63 mRNA and protein in in vitro cultivated promastigotes and that HE
XBP-deficient promastigotes were capable of lesion formation in BALB/c
mice.