EFFICIENCY OF INSERTION VERSUS REPLACEMENT VECTOR TARGETING VARIES ATDIFFERENT CHROMOSOMAL LOCI

We have analyzed the targeting frequencies and recombination products generated with isogenic vectors at the fah and fgr loci in embryonic s tem cells. A single vector which could be linearized at different site s to generate either a replacement or an insertion vector was construc ted for each locus. A replacement event predominated when the vectors were linearized at the edge of the homologous sequences, while an inse rtion event predominated when the vectors were linearized within the h omologous sequences. However, the ratio of the targeting frequencies e xhibited by the different vector configurations differed for the two-l oci. When the fgr vector was linearized as an insertion vector, the ra tio of targeted to random integrations was four- to eightfold greater than when the vector was linearized as a replacement vector. By contra st, the ratio of targeted to random integrations at the fah locus did not vary with the linearization site of the vector. The different rela tionships between the targeting frequency and the vector configuration at the fgr and fah loci may indicate a DNA sequence or chromatin stru cture preference for different targeting pathways.