Precise, Fast, Guaranteed.
KO mice by ingenious
Custom designs, speak with our scientists today.
ingenious began as one of the first pioneering companies delivering knockout mice. From conventional deletions to conditional knockouts and large scale deletions using BACs, we have developed the technology and expertise to target even the most challenging genes. Working with ingenious, our clients have comfort in knowing that they have chosen a company with longevity, backed by almost two decades of successful, published experience.
Utilizing both CRISPR and ES cell technologies, ingenious offers the following benefits
for producing your knockout mouse model:
As short as 6 months for germline-transmission knockout mouse models.
Assurance of your mouse model, not just your money back.
Utilizing our proprietary technologies.
- Targeted gene knockout mice with multiple options for strategy, including knockout with CRISPR/Cas9
- Delete as little as 200bp, or as much as 200kb using our BAC technology
- Enhance the versatility of your model by combining reporter gene expression with your targeted knockout
- Our experienced scientists listen, care, and deliver
Below are some of ingenious’core knockout specialties:
The target region to delete is replaced with a Neomycin selection cassette, or with a reporter gene. The targeted gene is thus inactivated at all times, in all tissues. Click to learn more.
The target region is flanked by loxP sites (“floxed”) to facilitate Cre recombinase-mediated excision, leading to tissue specific or temporal gene inactivation. Click to learn more.
No-Doubt Conditional Knockout™
Knockout your gene with certainty through GFP. By combining our split-GFP technology with our tried and true conditional targeting methodology, ingenious now offers the next generation of knockout mouse models. Our No-Doubt Conditional Knockout™ system provides strong GFP expression, independent of the endogenous gene promoter and in response to the Cre mediated recombination event, for straightforward identification of cells in which the gene deletion has occurred.
Large Scale Deletion
Base your designs around experiments, not limitations. Our proprietary BAC technology enables the specific deletion of very large genomic sequences, such as multiple genes or whole gene clusters, in a single targeting step. By innovating the design and construction of these once formidable targeting projects, ingenious has drastically reduced the timelines and costs associated with these knockout mouse models. At ingenious, 200 kb is the new standard.
For too long, mouse model designs have been dictated by size constraints imposed by the limitations of technology. At ingenious, our mission to push past limits has led to the development of proprietary BAC and fosmid technologies that reduce the time associated with these projects to a fraction of their typical imposing scale.
At ingenious 200 kb is the New Standard
From the initial project concept, to identifying positive clones, to confirming F1 neo deleted mice, all within a year- 200 kb targeting is the “standard size” at ingenious targeting laboratory. Design your model based on what is needed to advance your field, not on what someone else tells you is possible.
Gene Replacement with Neomycin Selection Cassette
A critical region of the target gene is replaced with the Neomycin selection cassette, which is also used for positive selection of targeted ES cell clones during tissue culture. As a result the gene is inactivated at all times, in all tissues. The Neo cassette can be retained in the mouse model to act as a potential gene-blocking “trap” within the gene locus that it was inserted into. Alternatively the cassette can be deleted via FLP recombination in vitro (using FLP ES cells) or in vivo (via mating to FLP transgenic mice). Because the Neo cassette can potentially disrupt other genes nearby, many researchers prefer to delete it.
Options for targeting strategies
- Delete all or most of the gene’s coding region – we are able to delete up to 200kb using large BACs
- Delete promoter related sequences together with at least the first coding exon, preferably more
- Delete an important domain without which the protein cannot function
- Aim to create a coding frameshift if only a portion of the gene is deleted
- No design is too difficult- ingenious’ experts can target even the most difficult genes, bringing your design ideas to life
Gene Replacement with Reporter
The target region can be replaced with a reporter gene, such that the reporter gene expression will be controlled by the target gene’s endogenous promoter. With this strategy a single mouse line combines gene knockout and visualization of that gene’s normal expression domain.
The reporter, including a polyA signal, is typically inserted at the endogenous ATG start site of the target gene of interest, usually also replacing the first exon of the gene. All other exons and introns can be kept intact to avoid deleting regulatory or promoter associated regions important for transcription from the target gene, for expressing the inserted reporter gene.
Some researchers prefer to not delete any endogenous gene sequence to avoid deleting important promoter related elements. In this case the reporter can be inserted at the ATG without replacing any gene sequence. The polyA signal included with the reporter gene should prevent read-through to avoid expression of the endogenous gene.
Contact us today to discuss ideas for your next constitutive knockout mouse.
Replace an entire genomic mouse gene with the human ortholog, including introns, in a single targeting step. Intronic sequences contain regulatory features, microRNA genes, promoters, enhancers, and splice sites for transcript isoforms. Ensure that you are working with the complete picture, because a gene is more than just the protein that it encodes.
“I am very happy with iTL’s mouse knockout services. It was pleasant to collaborate with my project manager. She was very thorough and communicative, an excellent combination. I will certainly use iTL again when needed, and I’m happy to share my positive experience with colleagues.”
-John Creemers, PhD