Targeting of exogenous DNA into Trypanosoma brucei requires a high degree of homology between donor and target DNA.
Integration of exogenous DNA into the trypanosome genome occurs by homologous recombination only. To test whether a high degree of homology between donor and target DNA is required, we have inserted marker genes for drug resistance into the promoter area of variant surface glycoprotein (VSG) gene expression sites of Trypanosoma brucei, using targeting fragments from two expression sites that are 92% identical. We observed integrations into expression sites that are known to be perfectly matched to the donor flanks, and into subsets of uncharacterized expression sites that are specific for each type of targeting fragment, and that could be similar or identical to the donor flanks. This requirement for very high homology was found in both procyclic and bloodstream-form trypanosomes. We speculate that trypanosomes have a mismatch repair system that suppresses recombination between divergent DNA sequences, and we discuss ways in which the trypanosome might circumvent the requirement for perfect DNA homology in the duplicative transposition of a VSG gene into a VSG gene expression site.
Blundell
P A
PA
Division of Molecular Biology, Netherlands Cancer Institute, Amsterdam.
Rudenko
G
G
Borst
P
P
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D016054:DNA, Protozoan; D004351:Drug Resistance; D016521:Electrophoresis, Gel, Pulsed-Field; D015870:Gene Expression; D005819:Genetic Markers; D006023:Glycoproteins / Q000235:genetics; D008969:Molecular Sequence Data; D011995:Recombination, Genetic; D012689:Sequence Homology, Nucleic Acid; D014170:Transformation, Genetic; D014346:Trypanosoma brucei brucei / Q000235:genetics*
D016054:DNA, Protozoan; D005819:Genetic Markers; D006023:Glycoproteins