UMMS Affiliation

Department of Cell Biology



Document Type


Medical Subject Headings

Animals; Blotting, Southern; Cloning, Molecular; Embryonal Carcinoma Stem Cells; *Genetic Vectors; Hypoxanthine Phosphoribosyltransferase; Mice; *Mutagenesis, Insertional; Neoplastic Stem Cells; Recombination, Genetic; Restriction Mapping


Cell Biology


Gene targeting has been used to direct mutations into specific chromosomal loci in murine embryonic stem (ES) cells. The altered locus can be studied in vivo with chimeras and, if the mutated cells contribute to the germ line, in their offspring. Although homologous recombination is the basis for the widely used gene targeting techniques, to date, the mechanism of homologous recombination between a vector and the chromosomal target in mammalian cells is essentially unknown. Here we look at the nature of gene targeting in ES cells by comparing an insertion vector with replacement vectors that target hprt. We found that the insertion vector targeted up to ninefold more frequently than a replacement vector with the same length of homologous sequence. We also observed that the majority of clones targeted with replacement vectors did not recombine as predicted. Analysis of the recombinant structures showed that the external heterologous sequences were often incorporated into the target locus. This observation can be explained by either single reciprocal recombination (vector insertion) of a recircularized vector or double reciprocal recombination/gene conversion (gene replacement) of a vector concatemer. Thus, single reciprocal recombination of an insertion vector occurs 92-fold more frequently than double reciprocal recombination of a replacement vector with crossover junctions on both the long and short arms.

Rights and Permissions

Citation: Mol Cell Biol. 1991 Sep;11(9):4509-17. Link to article on publisher's website

Related Resources

Link to Article in PubMed

Included in

Cell Biology Commons



To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.