Process of targeted mutagenesis in mice via homologous recombination in Embryonic Stem cells devides into the following steps:

Screening of libraries of 129/Ola genomic DNA and gene characterization

Construction of the targeting vector

Transfection of ES cells and clone picking

Screening of clones for identification of the desired targeted event

Selection, karyotyping and genotyping of ES cell clones

Injection of blastocysts and implantation into pseudopregnant foster mice

Production of pathogen free chimeras


In addition the following steps can be taken:

Test breeding to F1 generation

Production and verification of mutant homozygous mice

Back crossing onto C57BL/6 and/or 129/Ola genetic backgrounds