The present invention relates to providing methods for preparing a targeting construct for use in a targeting vector for gene targeting or homologous recombination. The invention also provides targeting vectors, and cells, plants and animals (including yeast) containing the vectors having predetermined modifications. The invention further provides plants and animals modified by the targeting vectors. The gene targeting methods used herein are based on transposon and recombination mediated procedures which provide for high throughput generation of deletions, which is amenable to seni automated production of knockout vectors.
