Starvax Service supplies custom knockout and knock-in transgenic services by utilizing our proprietary technology that is competitive and highly efficient for producing high quality gene targeting transgenics in embryonic stem cells (ES).

At Starvax Service we are committed to providing our clients with a high quality service. We understand that speed and reliability are crucial to ensuring client's success. We also ensure that our team is available to you at all times to promptly address your needs.
Gene targeting in mice involves several important steps:
Cloning & Mapping of Locus
Engineering a gene targeting construct
Generation of Targeted ES Cells
Production of Chimeric Mice
Germline transmission

Each of these steps requires close coordination between the investigators and Starvax Service. The services can be ordered together or individually depending on the investigator’s needs. The timeframe for completion of each step is relative and dependent upon the specific locus targeted for mutagenesis.

Cloning & Mapping of Locus to be Targeted

Starvax Service’s scientists work closely with clients during the initial screening and mapping procedures to ensure rapid and complete locus characterization. Comprehensive restriction maps are developed from multiple genomic clonal isolates to confirm nonrearrangement and mapping fidelity. Intron/exon borders as well as exonic coding sequence composition are confirmed by restriction mapping, sequencing and/or PCR..

Engineering a gene targeting construct

Once a genomic clone of the gene is in hand and restriction mapped, vector design and construction begins. The pRCH series of gene targeting vectors have been optimized for rapid and reliable gene targeting vector construction. These vectors are utilized for the customized construction of knockout or knockin vectors which allow for a high probability of gene functional inactivation. In addition, clients have the option to track the gene endogenous expression pattern through the construction of targeting vectors which contain reporters such as B-Galactosidase (LacZ) or luciferase.

Generation of Targeted ES Cells

Your construct will be transfected into ES cells by electroporation. Following successful electroporation, PPS knockout and knockin technologies provide the capacity for high throughput focused mutagenesis which results in successful gene targeting independent of genetic background and thus expands the possibilities for loci which may be mutated as well as cell types which may be custom engineered for genetic modifications. We therefore allow our clients a wide range of cell line choices not necessarily limited to murine embryonic stem cells. Gene targeting procedures are implemented in a rapid, streamlined fashion such that in most cases targeted events are identified within two weeks post transformation.

Production of Chimeric Mice

Upon the identification of targeted embryonic stem cell lines clonal cell stocks are thawed and expanded for blastocyst injection. Each stem cell line is reconfirmed for correct gene targeting by both Southern blot analysis and PCR genotyping. In addition, a number of standardized quality control parameters are implemented at this stage to guard against pre-injection stem cell differentiation. Independent injection experiments are performed which routinely result in the production of several chimeric mice.

Germline Transmission

In order to ensure germline transmission, several individual chimeric mice are intercrossed with a mouse strain chosen by the client for the production of heterozygotes. A number of different inbreeding as well as outbreeding strategies may be employed for the purposes of either enhancing or suppressing the phenotypic traits exhibited. It is at this point that heterozygotes may be transferred to the client or, for an additional fee, Starvax Service scientists will propagate the mice to homozygosity.