We have analyzed the targeting frequencies and recombination products generated with (1,(12)(13)(14)39). Vectors with a DSB in the homologous counterpart of the chromosomal sequences have been termed insertion vectors and usually recombine through a single reciprocal pathway (25).The DSB repair (DSBR) model, originally proposed to explain the repair of breaks in DNA, has been invoked to explain the elevation of targeting frequencies in S. cerevisiae observed by the provision of a DSB in the homologous sequences (24,27,35). Although this model was originally developed with S. cerevisiae, it is supported by data for vector-chromosome recombination in mammalian cells (12,14,39 (13, 41). These insertion-like products suggest a preference for the integration of replacement vectors via an insertion pathway after end-end joining, possibly via a mechanism similar to that described by the DSBR model (13, 38).The presence of free ends of a vector, either homologous or heterologous, may be important for directing specific targeted integration pathways which in turn may affect both the targeting frequencies and the integration products. Previously, it has been shown that at the Aprt locus in CHO cells and at an integrated simian virus 40 genome in COS1 cells, when a DSB is made in the homologous sequences of the vector, the gene targeting frequency increases relative to the frequencies observed when the DSB is made outside these elements (1,14). However, at the immunoglobulin heavy-chain constant region in hybridoma cells, replacement and insertion vectors target at about the same frequency (15). We have reported previously that at the hprt locus in embryonic stem (ES) cells insertion vectors target more efficiently than replacement vectors (11-13), yet the Capecchi laboratory (9, 37) has reported that the two vector types target at about the same efficiency.Free DNA ends are also important for extrachromosomal homologous recombination in mammalian cells. DSBs in the homologous sequences in either of the substrates involved in the exchange increase the recombination frequency (5, 6, 17). As predicted by the DSBR pathway, both insertion and replacement products are observed. However, other reports have shown that intermolecular extrachromosomal homologous recombination is increased if both introduced DNAs are linearized within or just close to the homologous sequences. To explain this observation, the single-strand annealing (SSA) model is invoked. In the SSA model, the breaks appear to be substrates for exonuclease or helicase activity which generates complementary single-stranded DNA ends. These homologous single-stranded regions can anneal to each other and result in a replacement product (2,19,30,40). The location of the break does not need to be flanked by homology as long as singlestrand homology is exposed after exonuclease digestion or 8385

A genetic linkage study, performed on a large family with autosomal dominant retinitis pigmentosa (RP), demonstrated that the RP gene may be linked to the Rh locus, known to be on the short arm of human chromosome 1. Linkage studies on RP along with other studies, can help to more accurately classify these disease entities. Localizing the RP gene locus has the potential for allowing the early diagnosis of individuals at risk.

During genetic linkage studies of retinitis pigmentosa in a Navajo Amerindian family, an apparent null allele of erythrocyte GPT (GPT⁰) was observed in a man and 2 of his daughters. This is the fourth description of a (GPT⁰) allele, the first outside of Europe and the first in the Navajo.

We have analyzed the targeting frequencies and recombination products generated with isogenic vectors at the fah and fgr loci in embryonic stem cells. A single vector which could be linearized at different sites to generate either a replacement or an insertion vector was constructed for each locus. A replacement event predominated when the vectors were linearized at the edge of the homologous sequences, while an insertion event predominated when the vectors were linearized within the homologous sequences. However, the ratio of the targeting frequencies exhibited by the different vector configurations differed for the two loci. When the fgr vector was linearized as an insertion vector, the ratio of targeted to random integrations was four- to eightfold greater than when the vector was linearized as a replacement vector. By contrast, the ratio of targeted to random integrations at the fah locus did not vary with the linearization site of the vector. The different relationships between the targeting frequency and the vector configuration at the fgr and fah loci may indicate a DNA sequence or chromatin structure preference for different targeting pathways.