title: Homologous recombination and stable transfection in the parasitic protozoan Trypanosoma brucei.
abstract: Development of methods for the manipulation of the genomes of parasitic protozoa will lead to enhanced understanding of parasite biology and host-parasite relationships. Efficient gene transfer and targeted integration by homologous recombination were achieved in the parasitic protozoan Trypanosoma brucei, the causative agent of sleeping sickness. An expression vector with the neomycin phosphotransferase gene (neo), under the control of a procyclic acidic repetitive protein (PARP) gene promoter, was targeted into an intergenic region in beta alpha-tubulin-gene tandem array. Sixteen copies of neo were found in a tandem array in one of the transfectants where the PARP promoter controlled alpha-amanitin-resistant transcription of neo, whereas transcription of tubulin genes remained alpha-amanitin-sensitive.
authors.affiliation: Division of Tropical Medicine, School of Public Health, Columbia University, New York, NY 10032.
authors.forename: M G
authors.initials: MG
authors.lastname: Lee
authors.forename: L H
authors.initials: LH
authors.lastname: Van der Ploeg
mesh_terms: D000546:Amanitins / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics; D004351:Drug Resistance / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D005839:Gentamicins; D019868:Kanamycin Kinase; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010770:Phosphotransferases / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000235:genetics; D015800:Protozoan Proteins; D011995:Recombination, Genetic; D015183:Restriction Mapping; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D014346:Trypanosoma brucei brucei / Q000235:genetics*; D014404:Tubulin / Q000235:genetics; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000235:genetics
chemical_list: D000546:Amanitins; D016054:DNA, Protozoan; D005839:Gentamicins; D008562:Membrane Glycoproteins; D015800:Protozoan Proteins; D014404:Tubulin; D014643:Variant Surface Glycoproteins, Trypanosoma; C053277:procyclic acidic repetitive protein, Trypanosoma; C010680:antibiotic G 418; D015500:Chloramphenicol O-Acetyltransferase; D010770:Phosphotransferases; D019868:Kanamycin Kinase