Knockin Approaches
Point Mutation Knockin
Point mutation knockin models introduce specific nucleotide changes into the endogenous gene. This approach is essential for modeling human disease alleles, studying structure function relationships, and creating drug resistant or drug sensitized variants for pharmacological studies.
Point mutations can be introduced as constitutive changes present from conception, or as conditional alleles that convert from wildtype to mutant sequence upon Cre mediated recombination.
Learn more→Reporter Knockin
Reporter knockin models insert visualization markers at endogenous loci. Common reporters include fluorescent proteins (GFP, tdTomato, mCherry), enzymatic reporters (LacZ, alkaline phosphatase), and bioluminescent reporters (luciferase). Reporter expression is driven by the endogenous promoter, providing accurate readout of native gene expression patterns.
Reporter knockins enable lineage tracing, cell sorting of expressing populations, and real time monitoring of gene expression in living animals.
Learn more→Tag Knockin
Tag knockin models add epitope tags to endogenous proteins for biochemical detection, purification, or localization studies. Common tags include FLAG, HA, V5, and Myc. Tags can be positioned at the N terminus, C terminus, or internal locations depending on protein structure and experimental requirements.
Because tagged proteins are expressed at physiological levels under native regulatory control, tag knockin models avoid overexpression artifacts common with transgenic approaches.
Learn more→cDNA Knockin
cDNA knockin models replace the endogenous coding sequence with an alternative cDNA. Applications include humanization (replacing mouse gene with human ortholog), isoform specific expression, and introduction of modified or optimized coding sequences.
cDNA knockins maintain endogenous regulatory elements while providing complete control over the expressed protein sequence.
Learn more→Conditional Knockin
Conditional knockin models express modified alleles only after Cre mediated activation. A stop cassette flanked by LoxP sites prevents expression until Cre excision. This approach enables tissue specific or temporally controlled expression of mutant alleles, reporters, or other modifications.
Conditional knockins are particularly valuable for studying oncogenic mutations that would be lethal if expressed globally from conception.
Learn more→Technical Approach
Allele Design
Knockin allele design requires careful consideration of the modification site, surrounding sequence context, and potential effects on gene expression. Our scientific team analyzes:
- Optimal insertion site for reporters and tags
- Codon context for point mutations
- Splice site preservation
- Regulatory element maintenance
When to Use Knockin Models
Knockin models provide the most physiologically relevant approach when native gene regulation is important for the experimental question.
Knockin vs Alternative Approaches
| Research Goal | Knockin Approach | Alternative |
|---|---|---|
| Model human disease mutation | Point mutation knockin | Transgenic (but loses native regulation) |
| Visualize gene expression | Reporter knockin | Transgenic reporter (but random integration) |
| Track protein localization | Tag knockin | Antibody (but requires fixation) |
| Express human protein | cDNA knockin / Humanization | Transgenic (but overexpression artifacts) |
| Tissue specific mutant expression | Conditional knockin | Viral delivery (but variable efficiency) |
Selected Publications
Knockin models generated by ingenious targeting laboratory:
Diamond EL, Emile JF, Fujino T, Haroche J, Maron MI, Lewis AM, Rahman J, Reiner AS, Bossert D, Rosenblum M, Yabe M, Petrova-Drus K, Francis JH, Rotemberg V, Rampal RK, Yoo S, Daniyan AF, Mahajan S, Hatzoglou V, Young R, Ulaner GA, Rösler W, Hershkovitz-Rokah O, Shpilberg O, Mazor RD, Chen LYC, Singer M, Cuibus MA, Weis K, Benbarche S, Zhang P, Fox N, Castro C, Tittley S, Witkowski M, Cohen-Aubart F, Terriou L, Hanoun M, Schleinitz N, Sosa G, Hautala T, De Lassus LF, Rosen N, Abdel-Wahab O, Durham BH. (2025).
RAF-independent MEK mutations drive refractory histiocytic neoplasms but respond to ERK inhibition ↗Cancer Cell Online ahead of print
Jiang Y, Sachdeva K, Goulbourne CN, Berg MJ, Peddy J, Stavrides PH, Pensalfini A, Pawlik M, Malampati S, Whyte L, Basavarajappa BS, Shivakumar S, Bleiwas C, Smiley JF, Mathews PM, Nixon RA. (2025).
Increased neuronal expression of the early endosomal adaptor APPL1 leads to endosomal and synaptic dysfunction with cholinergic neurodegeneration ↗J Neurosci 29(45): e2331242025
Mohassel P, Hearn H, Rooney J, Zou Y, Johnson K, Norato G, Nalls MA, Yun P, Ogata T, Silverstein S, Sleboda DA, Roberts TJ, Rifkin DB, Bönnemann CG. (2025).
Collagen type VI regulates TGF-β bioavailability in skeletal muscle in mice ↗J Clin Invest 9(135): e173354
What Researchers Say
“The people at InGenious are friendly, professional, and extremely good at what they do. I have made 5 Knockin mice with them and everything has gone like clockwork.”
— David B. Roth, MD, PhD
Perelman School of Medicine, University of Pennsylvania
Start Your Knockin Project
Our scientific consultants are ready to discuss your research requirements and recommend the optimal knockin strategy for your experimental goals. Initial consultation is provided at no charge and includes modification site analysis, allele design recommendations, and timeline estimates.
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Key Terms
Understanding the terminology used in knockin mouse model generation helps you communicate effectively with our scientific team and interpret project documentation.
Knockin
A gene targeting strategy that inserts a specific DNA sequence at a defined genomic location, such as reporter genes, point mutations, or human sequences.
Reporter Gene
A gene encoding an easily detectable protein used to visualize gene expression patterns. Common reporters include fluorescent proteins and LacZ.
Point Mutation
A change in a single nucleotide within a gene sequence. Point mutation knockin models introduce specific mutations to study disease mechanisms.
Safe Harbor Locus
A genomic location where transgene insertion does not disrupt essential gene functions and supports stable expression.