Gene Targeting
Would you like to explore more biology or learn more about your gene/target of interest? Creating and characterizing a gene knockout mouse is the most well accepted strategy for evaluating gene function. The knockout mouse is one that has been genetically engineered such that one or more of functions of a gene(s) has made “null” through a gene ablation process involving homologous recombination techniques. By inactivating the gene and studying the mouse for any resulting differences, researchers can infer the probable function(s) associated with that gene. For more information regarding the characterization of such knockout or knockin mouse lines, please visit our page describing CDAS capabilities in “Comprehensive/Therapeutic Area Focused Phenotyping”.The creation of standard or conditional knockout mice, or knockin mice, utilizes some proprietary gene targeting methodologies developed in our state-of-the-art facilities. Through these gene modification processes, CDAS is able to produce a customized collection of genetic antagonists, including conventional or conditional knockouts/knockins, each capable of possessing spatial- and temporal-specific modifications. Our Gene Targeting packages are customizable, enabling customers to select from multiple entry points, such as (i) gene sequence or accession number, (ii) transmitted targeting vector, or (iii) third party ES cell clone(s).
Following the creation of a customized targeting vector, the electroporation step utilizes our highest quality germline-competent mouse embryonic stem (ES) cells – either the popular C57BL/6, 129/SvJ, 129/SvEv, or FVB/N. The transfected ES cells are then isolated using positive/negative selection and Southern blot analysis techniques. Finally, targeted ES cell clones are injected into mouse recipient blastocysts in order to produce high percentage chimeric animals. Generally, our processes and targeting vectors incorporate the ability to remove the Neo (or puromycin) selection cassette in cell culture.
Whether the goals are customized gene targeted animal models including standard knockouts or knockins, or conditional knockouts or knock-ins of known genes, a gene family, mutant genes or artificially created reporters and/or gene constructs we have all the tools and experience to deliver a higher probability of success. CDAS has more than 18 years’ worth of experience in using ES cell technology – moreover, we have designed, produced, and characterized over 800 knockout/knockin lines while maintaining a 97% overall success rate. Ultimately, we have isolated the most robust and dependable ES cell lines in the industry.
For simplicity, our complete package involves cloning and characterization of mouse genomic target DNA, design and construction of a specific targeting vector, transfection of proprietary ES cells, screening of up to 200 individual drug resistant clones, injection of up to two (2) targeted ES clones into recipient blastocysts, generation of high-percentage chimeras from injected blastocysts, and breeding of chimera to F1 generation heterozygotic animals. While offering entry point flexibility at the outset of the project, CDAS also offers optional capabilities such as breeding to hetero/homozygosity, and removal of selection cassettes in tissue culture.
For more information please review the related downloads, email us directly at CDASinvivo@caliperls.com or complete and submit the product inquiry form and a CDAS representative will contact you.
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