Episomal and integrated maintenance of foreign DNA in XXXX .

@entity461 @entity1412 @entity344 @entity5808
@entity461 is an early diverging eukaryote which causes @entity344 throughout the world. Different subgroups of Giardia have been defined based on several biochemical and genetic criteria. We have developed a method for stably introducing DNA into the nuclei of the parasite using @entity5808 acetyltransferase (pac) as a dominant selectable marker. Transfected circular DNAs were maintained as episomes in the isolate WB, a representative of one Giardia subgroup. When input DNAs were linearized, integration was observed to occur by homologous recombination producing gene replacements in this isolate. In isolate @entity1412 , which represents a different subgroup, both linear and circular transfected DNAs were integrated into the genome by homologous recombination. In @entity1412 , linear DNA again produced gene replacements, while circular DNA produced duplicative integration events. The failure of @entity1412 to replicate episomes may reflect differences in the structure or recognition of DNA replication origins between these subgroups. A plasmid shuttle vector was also developed for expression of other genes in @entity461 . Utilizing the green fluorescent protein as a reporter gene in the WB isolate, we show that gene expression from this vector correlated with plasmid copy number over a range of two orders of magnitude. Together these tools should greatly enhance our ability to study both the basic biology and the pathogenesis of this ubiquitous parasite.
