Point Mutation Mouse Models

Precision targeting for your research.

A point mutation model from ingenious allows you to study a known disease-causing mutation or alter a critical site from the murine to human sequence. Different strategies are available depending on the experiments you have planned. No design is out of reach – our experienced staff will work with you to identify the best options for your gene of interest.

Conventional and Conditional Point Mutation Knockin Approaches

Conventional Point Mutation

A conventional point mutation results in the targeted allele encoding only the mutant sequence. This can be a powerful model for human disease, for example by changing a crucial codon to cause a deleterious amino acid substitution. If you want more control over expression of the mutant sequence consider a conditional point mutation strategy, described below.

Conditional Point Mutation

Conditional point mutant lines don’t express the mutant sequence until you choose to activate it. Your tissue-specific Cre-expressing lines allow expression of the mutant gene only where Cre is active. Depending on your needs the allele can initially express the wild-type sequence or function as a null. Bypass the deleterious effects of your mutation on embryonic development and create a model where you control the timing and location of activation.

Wildtype-First with Conditional Activation of Mutant

Advances in molecular biology and gene targeting now allow for generating animal models with the ability to “turn on” a mutation in a specific tissue and/or at a specific time point. The gene will express as wild-type initially, until Cre recombinase is activated. Cre deletes the wild-type sequence and allows expression of the mutant sequence.

There is a lower risk of embryonic lethality or artificial phenotype since the mutation would be activated in only a specific tissue, or at a specific time point. Furthermore, a tissue or time specific mutation may more accurately reflect the disease of study. With the variety of Cre transgenic mouse lines available, multiple lines can be generated from the parental targeted mouse line, for expressing the mutant in different tissues or at specific times (CreERT2 or tetO-Cre can also be used for this). Thus the mouse model is more versatile and can be utilized across scientific disciplines.

The following are two design examples on how this can be accomplished:

Minigene Approach

Notable Client Publications

1) Cortes JR, Ambesi-Impiombato A, Couronne L, Quinn SA, Kim CS, et al. 2018. RHOA G17V Induces T Follicular Helper Cell Specification and Promotes Lymphomagenesis. Cancer Cell 33(2): 259-273.

2) Beguelin W, Teater M, Gearhart MD, Calvo Fernandez MT, Goldstein RL, et al. 2016. EZH2 and BCL6 Cooperate to Assemble CBX8-BCOR Complex to Repress Bivalent Promoters, Mediate Germinal Center Formation and Lymphomagenesis. Cancer Cell 30: 197-213.

3) Kim E, Ilagan JO, Liang Y, Daubner GM, Lee SC, et al. 2015. SRSF2 Mutations Contribute to Myelodysplasia by Mutant-Specific Effects on Exon Recognition. Cancer Cell 27(5): 617-630.

Inversion Approach

Notable Client Publications

1) Venturutti L, Teater M, Zhai A, Chadburn A, Babiker L, et al. 2020. TBL1XR1 Mutations Drive Extranodal Lymphoma by Inducing a Pro-tumorigenic Memory Fate. Cell 182(2): 297-316.e27.

Knockout-First with Conditional Activation of Mutant

In some cases, our clients require more sophisticated mutation design options to allow for inducible expression of their mutation. One method that has been very successful and widely adapted is the use of removable transcription termination elements to control the expression of targeted alleles. Excision of the STOP cassette, and resulting expression of the mutant gene, is mediated by inducible and/or lineage specific recombinases (e.g., Cre or FLP), which allows spatial and temporal control of gene expression.