Formation of extrachromosomal circular amplicons with direct or inverted duplications in drug-resistant Leishmania tarentolae.
Selection for methotrexate resistance in Leishmania spp. is often associated with amplification of the H locus short-chain dehydrogenase-reductase gene ptr1 as part of extrachromosomal elements. Extensive sequences are always coamplified and often contain inverted duplications, most likely formed by the annealing of inverted repeats present at the H locus. By gene targeting mediated by homologous recombination, several repeated sequences were introduced in the vicinity of ptr1. Selection for methotrexate resistance in these transfectants led to ptr1 amplification as part of small circles with direct or inverted duplications whether the integrated sequences consisted of direct or inverted repeats. Hence, for a region to he amplified in L. tarentolae during drug selection, a drug resistance gene is required and must be flanked by (any) homologous repeated sequences. The distance between these repeats and their orientation will determine the length of the amplicon and whether it contains direct or inverted duplications.
Grondin
K
K
Centre de Recherche en Infectiologie du Centre Hospitalier de l'Université Laval, Québec, Canada.
Roy
G
G
Ouellette
M
M
D000818:Animals; D002460:Cell Line; D016054:DNA, Protozoan / Q000378:metabolism; D004351:Drug Resistance / Q000235:genetics*; D018274:Electroporation; D005784:Gene Amplification; D015321:Gene Rearrangement; D017125:Genes, Protozoan; D007891:Leishmania / Q000187:drug effects / Q000235:genetics*; D008727:Methotrexate / Q000494:pharmacology*; D008957:Models, Genetic; D010088:Oxidoreductases / Q000235:genetics*; D011995:Recombination, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D014162:Transfection
D016054:DNA, Protozoan; D010088:Oxidoreductases; C085592:pteridine reductase; D008727:Methotrexate