Patent ID: 6897353
Filing Date: 2005-05-24
Classification: C12N,C12Q

Abstract:
1. A method of producing a transgenic, non-human mammal having integrated into its genome a defined segment of DNA, comprising: a. obtaining a yeast-bacteria shuttle vector comprising a yeast replication origin, a yeast selection marker gene, a bacterial replication origin, a bacterial selection marker gene, at least one unique cloning site, and sequences homologous to sequences flanking the defined segment of DNA, wherein said bacterial replication origin is selected from the group consisting of P1 replicon and F factor origin of replication; b. linearizing the vector within the homologous sequences to form recombinogenic ends; c. introducing the linearized vector into a yeast cell containing DNA comprising the defined segment of DNA, wherein the linearized vector and defined segment of DNA homologously recombine to form a recombinant product; d. selecting for the recombinant product; e. transferring the recombinant product to bacteria for amplification; f. isolating the defined segment of DNA from the recombinant product; g. integrating the defined segment of DNA into the genome of a mammal by microinjection of a fertilized egg; h. transferring the fertilized egg containing the defined segment of DNA to a pseudopregnant non-human mammal; and i. allowing the fertilized egg containing the defined segment of DNA to develop into a transgenic, non-human mammal.