Patent ID: 6503712
Filing Date: 2003-01-07
Classification: A01K,C12N

Abstract:
A method of preparing a genomic library for use in producing knockout targeting vectors comprising:a) preparing genomic DNA fragments of pre-selected sizes wherein said genomic DNA comprises mouse genomic DNA fragments ranging from 8 kb to 14 kb; b) preparing a shuttle vector comprising inserting said genomic DNA fragments into a yeast vector wherein said yeast vector is pYYL-1, which comprises: i) a bacterial origin of replication; ii) a bacterial selection marker; iii) a yeast origin of replication; iv) a yeast selection marker; and v) a selectable marker for selection of mammalian cells; c) introducing said vector into bacterial host cells to amplify said shuttle vectors in transformed bacterial host cells; and; d) arraying said transformed bacterial host cells into pools wherein the bacterial host cells comprise said shuttle vectors and wherein said pools comprise genomic fragments of pre-selected sizes wherein said pYYL-1 vector comprises a 4.4 kb fragment, generated through a SphI restriction digestion of yeast-E. coli shuttle vector pGBT9, wherein said 4.4 kb fragment is annealed at the SphI sites to a first oligonucleotide containing a SP6 site and a second oligonucleotide containing a T7 site.