# Knockout Mouse: A Comprehensive Guide to Gene Function Studies

## Introduction to Knockout Mice

### What is a Knockout Mouse?

A knockout mouse is a genetically engineered laboratory mouse in which a specific gene has been inactivated or "knocked out" through targeted modification of its genome. The term "knockout" refers to the complete disruption of gene function, typically achieved by deleting a critical exon, inserting a premature stop codon, or introducing a [frameshift mutation](/knowledge/molecular-biology/frameshift-mutation) that renders the gene product non-functional. Because the mouse genome shares approximately 85% of its protein-coding genes with the human genome, knockout mice serve as powerful experimental models for determining what a gene does in a living organism.

The fundamental principle underlying knockout technology is gene targeting—the deliberate modification of a specific, predetermined locus in the genome. This is distinct from random mutagenesis approaches, where mutations are introduced indiscriminately and the investigator must screen for a phenotype of interest. Gene targeting allows researchers to ask a precise question: what happens when this particular gene is non-functional?

### Why Are Knockout Mice Important?

Knockout mice are important because they provide a direct, causal link between genotype and phenotype. While cell culture systems can reveal a gene's function in isolation, they cannot capture the complexity of a whole organism—the interplay between tissues, the effects of developmental timing, and the physiological responses that only emerge at the organismal level. For example, knocking out the *leptin* gene (*ob*) in mice produces profound obesity, hyperphagia, and diabetes, revealing the gene's role in appetite regulation and energy homeostasis. This finding directly informed the discovery of leptin's function in humans and led to therapeutic approaches for certain forms of obesity.

Knockout mice are also essential for validating drug targets. If a gene is proposed as a therapeutic target, a knockout mouse can reveal whether its absence causes unacceptable toxicity or whether it produces a phenotype that supports the therapeutic hypothesis. The [Knockout Mice](/knowledge/molecular-biology/knockout-mice) resource at the International Mouse Phenotyping Consortium has generated thousands of knockout lines, providing a systematic view of gene function across the genome.

## History and Development

### Early Gene Targeting in Mice

The conceptual foundation for gene targeting emerged in the 1970s with the discovery of [homologous recombination](/knowledge/molecular-biology/homologous-recombination)—the cellular process by which DNA sequences with high similarity exchange genetic information. In 1977, Mario Capecchi demonstrated that exogenous DNA introduced into mammalian cells could integrate into the genome by [homologous recombination](/knowledge/molecular-biology/homologous-recombination), albeit at a very low frequency. The challenge was to make this process efficient enough to be practical.

The key breakthrough came from the use of embryonic stem (ES) cells, which are pluripotent cells derived from the inner cell mass of a blastocyst-stage embryo. In 1981, Martin Evans and Matthew Kaufman independently isolated mouse ES cells and showed they could be maintained in culture while retaining the ability to contribute to all tissues of a developing embryo, including the germline. This property was essential: it meant that genetic modifications made in cultured ES cells could be transmitted to subsequent generations of mice.

### The First Knockout Mouse

In 1989, three independent groups—Mario Capecchi, Oliver Smithies, and Martin Evans—reported the first successful generation of knockout mice. The first gene targeted was *Hprt* (hypoxanthine-guanine phosphoribosyltransferase), chosen because it could be selected for in culture using the drug 6-thioguanine. Cells lacking functional HPRT survive in the presence of this drug, while cells with functional HPRT die. This positive-negative selection strategy allowed the researchers to enrich for ES cells that had undergone the desired homologous recombination event.

The achievement was recognized with the 2007 Nobel Prize in Physiology or Medicine, awarded to Capecchi, Smithies, and Evans. Their work established the blueprint for gene targeting that would dominate the field for the next two decades.

### Evolution of Techniques

The original gene targeting approach relied on homologous recombination in ES cells, a process with an efficiency of roughly 1 in 10⁶ to 10⁷ cells. The introduction of positive-negative selection using neomycin resistance (*neo*) and thymidine kinase (*tk*) cassettes improved enrichment but remained labor-intensive. A single knockout mouse line typically required 1–2 years to generate.

The field was revolutionized in 2013 with the application of CRISPR/Cas9 technology to mouse embryos. The CRISPR system, derived from a bacterial adaptive immune system, uses a guide RNA to direct the [Cas9 endonuclease](/knowledge/bioinformatics/genes/microbiology-amr/cas9-gene-structure-function-pathway) to a specific genomic sequence, where it introduces a double-strand break. The cell's error-prone non-homologous end joining (NHEJ) repair pathway then introduces insertions or deletions (indels) at the cut site, frequently causing frameshift mutations that knock out gene function. This approach can be performed directly in zygotes, bypassing ES cell culture entirely, and can generate knockout mice in as little as 6–8 weeks. For a detailed comparison of methods, see [CRISPR Knockout](/knowledge/molecular-biology/crispr-knockout).

## How Knockout Mice Are Created

### Traditional Gene Targeting via Homologous Recombination

The traditional method for generating knockout mice involves several sequential steps:

1. **Design and construction of the targeting vector.** A targeting vector is a plasmid containing a modified version of the gene of interest. The vector includes: (a) a neomycin resistance cassette (*neo*) that replaces a critical exon or the entire coding sequence, (b) 5' and 3' homology arms—sequences of 5–10 kilobases that match the genomic locus flanking the region to be deleted, and (c) a thymidine kinase (*tk*) cassette outside the homology arms for negative selection.

2. **Electroporation of ES cells.** The linearized targeting vector is introduced into mouse ES cells by electroporation. A typical electroporation uses 10–25 µg of DNA for 10⁷ cells in a 0.4 cm cuvette at 250 V and 500 µF. The cells are then subjected to selection with G418 (geneticin) at 200–400 µg/mL to select for integration of the *neo* cassette, and ganciclovir (2 µM) to select against random integration events that retain the *tk* cassette.

3. **Screening for homologous recombination.** Surviving colonies are expanded and screened by Southern blotting or PCR to identify clones where the targeting vector integrated at the correct locus. The frequency of correctly targeted clones typically ranges from 1% to 20% of surviving colonies, depending on the locus.

4. **Blastocyst injection or morula aggregation.** Correctly targeted ES cells are injected into blastocysts (3.5 days post-coitum) or aggregated with morulae (2.5 days post-coitum). The injected blastocysts are then transferred into pseudopregnant foster mothers.

### CRISPR/Cas9-Mediated Knockout

The CRISPR/Cas9 approach is faster and more flexible than traditional gene targeting:

1. **Guide RNA design.** A single-guide RNA (sgRNA) of approximately 20 nucleotides is designed to be complementary to the target sequence, immediately upstream of a protospacer adjacent motif (PAM) sequence (NGG for *Streptococcus pyogenes* Cas9). The sgRNA and Cas9 mRNA or protein are co-injected into the cytoplasm of fertilized mouse zygotes.

2. **Microinjection into zygotes.** Injection is performed using a micromanipulator under a microscope. A typical injection mixture contains 50 ng/µL Cas9 mRNA and 25 ng/µL sgRNA in injection buffer (10 mM Tris-HCl, pH 7.5, 0.1 mM EDTA). Alternatively, Cas9 protein complexed with sgRNA (ribonucleoprotein, RNP) can be injected at 30 ng/µL.

3. **Embryo transfer.** Injected zygotes are cultured briefly and transferred into the oviducts of pseudopregnant females. Approximately 50–70% of injected zygotes develop to term.

4. **Genotyping.** Founder mice (F0) are genotyped by PCR amplification of the target region followed by sequencing or [restriction enzyme digestion](/knowledge/diagnostics/molecular/restriction-enzyme-digestion-protocol-troubleshooting). Mice carrying frameshift mutations in the target gene are selected for breeding.

### Generating Chimeric Mice and Breeding

In the traditional ES cell method, the blastocysts injected with targeted ES cells produce chimeric mice—animals composed of cells derived from both the host blastocyst and the injected ES cells. The extent of chimerism is assessed by coat color: if the ES cells are derived from a mouse strain with a different coat color (e.g., 129/Sv, agouti) than the host blastocyst (e.g., C57BL/6, black), the degree of agouti contribution indicates the proportion of ES cell-derived tissue.

Chimeric males are bred to wild-type females, and germline transmission is confirmed when the agouti coat color appears in the offspring. Heterozygous offspring (carrying one knockout allele) are then intercrossed to generate homozygous knockout mice. The breeding scheme follows Mendelian genetics: a heterozygous × heterozygous cross yields 25% wild-type, 50% heterozygous, and 25% homozygous knockout offspring.

## Types of Knockout Mice

### Constitutive Knockouts

A constitutive knockout has the gene deleted in all cells and tissues from the earliest stages of development. This is the default outcome of both traditional gene targeting and CRISPR-mediated knockout. Constitutive knockouts are valuable for determining the overall function of a gene, but they have a significant limitation: if the gene is essential for development, the knockout may be embryonic lethal, preventing analysis of the gene's function in adult tissues.

For example, knockout of the *Brca1* gene, involved in DNA repair, results in embryonic lethality at approximately day 7.5 of gestation due to proliferation defects. While this demonstrates that *Brca1* is essential for early development, it precludes studying its role in adult tissues such as the mammary gland, where it functions as a tumor suppressor.

### Conditional Knockouts (Cre-loxP)

Conditional knockouts allow gene deletion to be restricted to specific tissues or cell types. The most widely used system is the Cre-loxP system, derived from bacteriophage P1. The Cre recombinase recognizes 34-base pair loxP sites and catalyzes recombination between them, deleting any DNA sequence flanked by two loxP sites in the same orientation.

To generate a conditional knockout, two mouse lines are required:

1. **The floxed line.** The target gene is flanked by loxP sites ("floxed") through homologous recombination in ES cells. The loxP sites are typically inserted into introns, leaving the gene fully functional.

2. **The Cre line.** A transgenic mouse line expressing Cre recombinase under the control of a tissue-specific or cell-type-specific promoter.

When the two lines are crossed, offspring carrying both the floxed allele and the Cre transgene will have the gene deleted only in cells where the promoter driving Cre is active. For example, crossing a floxed *Trp53* (p53) mouse with a mouse expressing Cre under the control of the *MMTV* promoter (active in mammary epithelium) produces mice with p53 deleted specifically in mammary tissue, allowing study of p53's role in breast cancer without the systemic effects of global deletion.

### Inducible Knockouts

Inducible knockouts add a temporal dimension to conditional deletion. The most common system uses a fusion protein of Cre and a modified estrogen receptor ligand-binding domain (CreERT2). This fusion protein is retained in the cytoplasm until the synthetic ligand tamoxifen (or its metabolite 4-hydroxytamoxifen) is administered. Tamoxifen binds to the estrogen receptor domain, allowing Cre to translocate to the nucleus and catalyze recombination.

A typical induction protocol involves intraperitoneal injection of tamoxifen (75–100 mg/kg body weight) for 5 consecutive days. Gene deletion occurs within 2–5 days after the first injection. This system allows researchers to delete a gene in adult animals, avoiding developmental compensation and embryonic lethality. For example, inducible knockout of the *Apc* gene in adult intestinal epithelium leads to rapid formation of intestinal adenomas, modeling familial adenomatous polyposis.

## Applications of Knockout Mice

### Gene Function Analysis

The most fundamental application of knockout mice is determining what a gene does. By comparing knockout animals to wild-type littermates across a range of phenotypes, researchers can infer the gene's function. The International Mouse Phenotyping Consortium (IMPC) has established a standardized phenotyping pipeline that assesses over 250 parameters, including body weight, blood chemistry, behavior, immune function, and histopathology.

For example, knockout of the *Clock* gene, a core component of the circadian clock, produces mice with disrupted circadian rhythms, reduced sleep, and metabolic abnormalities. This establishes *Clock* as a master regulator of circadian behavior and metabolism.

### Disease Models

Knockout mice are invaluable for modeling human genetic diseases. Mice with mutations in genes orthologous to those causing human diseases recapitulate many aspects of the human condition, providing platforms for understanding disease mechanisms and testing therapies.

Notable examples include:

- **Cystic fibrosis:** Knockout of *Cftr* produces mice with defective chloride transport, although the lung phenotype is milder than in humans.
- **Duchenne muscular dystrophy:** The *mdx* mouse, which carries a spontaneous mutation in the *Dmd* gene, and engineered knockouts show muscle degeneration and weakness.
- **Neurodegenerative diseases:** Knockout of *Park2* (parkin) or *Pink1* produces mice with mitochondrial dysfunction and subtle motor deficits, modeling aspects of Parkinson's disease.
- **Cancer:** Knockout of [tumor suppressor genes](/knowledge/molecular-biology/tumor-suppressor-gene) such as *Trp53*, *Rb1*, and *Apc* produces mice with increased tumor susceptibility, providing platforms for testing chemopreventive agents.

### Drug Testing and Development

Knockout mice are used throughout the drug development pipeline. Target validation is a critical early step: if a drug is designed to inhibit a specific protein, a knockout mouse can reveal whether loss of that protein's function produces a phenotype consistent with the desired therapeutic effect. Knockout mice are also used to assess on-target toxicity—side effects that arise from inhibiting the intended target rather than from off-target interactions.

For example, before developing PCSK9 inhibitors for hypercholesterolemia, researchers demonstrated that *Pcsk9* knockout mice have dramatically reduced plasma cholesterol levels and are protected from atherosclerosis. This provided strong validation for PCSK9 as a drug target, and the subsequent inhibitors (evolocumab, alirocumab) are now approved for clinical use.

## Phenotypic Analysis of Knockout Mice

### Behavioral Tests

Behavioral phenotyping is a critical component of knockout mouse analysis, particularly for genes implicated in neurological and psychiatric disorders. Standardized tests include:

- **Open field test:** Measures locomotor activity and anxiety-like behavior by tracking movement in a novel arena over 5–10 minutes.
- **Elevated plus maze:** Assesses anxiety by measuring time spent in open versus closed arms over 5 minutes.
- **Morris water maze:** Tests spatial learning and memory. Mice are trained to find a hidden platform in a pool of opaque water over 5–10 days, with probe trials assessing memory retention.
- **Tail suspension test and forced swim test:** Measure behavioral despair, used as screens for antidepressant-like effects.

### Molecular and Cellular Assays

Molecular analyses confirm the knockout at the DNA, RNA, and protein levels and assess downstream consequences:

- **Genotyping:** PCR amplification of the target locus confirms the presence of the knockout allele.
- **Quantitative RT-PCR (qRT-PCR):** Measures mRNA expression levels. A complete knockout should show no detectable transcript, although some truncated transcripts may persist.
- **Western blotting:** Confirms absence of the protein product. This is essential because some knockout strategies produce truncated proteins that retain partial function.
- **Enzyme activity assays:** For enzymes, direct measurement of catalytic activity provides functional confirmation of knockout.

### Histological Examination

Histopathology reveals structural abnormalities in tissues. Standard procedures involve:

1. **Tissue collection and fixation.** Tissues are collected and fixed in 10% neutral-buffered formalin for 24–48 hours at room temperature.
2. **Embedding and sectioning.** Fixed tissues are embedded in paraffin and sectioned at 4–6 µm thickness using a microtome.
3. **Staining.** Hematoxylin and eosin (H&E) staining is the standard for general morphology. Specialized stains include Masson's trichrome for collagen, Oil Red O for lipids, and immunohistochemistry for specific proteins.

For example, in a knockout mouse model of atherosclerosis, histological examination of the aortic root with Oil Red O staining reveals lipid accumulation in the vessel wall, while immunostaining for macrophage markers (e.g., CD68) identifies inflammatory infiltrates.

## Limitations and Challenges

### Embryonic Lethality

Many genes are essential for development, and constitutive knockout results in embryonic or perinatal lethality. This limits analysis to early developmental stages and prevents study of the gene's function in adult tissues. The *Rb1* (retinoblastoma) knockout, for example, dies at embryonic day 13.5 with defects in hematopoiesis and neuronal development. Conditional and inducible knockout strategies are the primary solutions to this problem, allowing gene deletion at later time points or in specific tissues.

### Genetic Redundancy

Gene families often contain members with overlapping functions. Knocking out one gene may produce no phenotype because a paralog compensates for its loss. For example, knockout of *MyoD* alone produces viable mice with apparently normal skeletal muscle, because the related factor Myf5 compensates during development. Only when both *MyoD* and *Myf5* are knocked out does muscle development fail completely. This redundancy can mask the true function of a gene and requires the generation of double or triple knockouts to reveal phenotypes.

### Off-Target Effects

CRISPR/Cas9 can introduce mutations at sites other than the intended target, particularly if the guide RNA shares sequence similarity with other genomic loci. Off-target effects can confound phenotypic analysis by introducing additional mutations that contribute to the observed phenotype. Mitigation strategies include:

- Using guide RNAs with minimal predicted off-target sites (verified by computational tools such as Cas-OFFinder).
- Using high-fidelity Cas9 variants (e.g., eSpCas9, HiFi Cas9) that reduce off-target cleavage.
- Outcrossing founder mice to wild-type animals for at least two generations to segregate away off-target mutations.

### Background Strain Effects

The genetic background of the mouse strain significantly influences phenotype. A knockout that produces a severe phenotype on one background may be mild or undetectable on another. For example, *Apc* mutations cause intestinal polyposis on the C57BL/6 background but are less penetrant on other backgrounds due to modifier genes. Researchers must therefore consider the strain background when interpreting phenotypes and when comparing results across studies. Standard practice is to backcross knockout lines to a defined strain (typically C57BL/6) for at least 10 generations to achieve >99% congenic status.

## Ethical Considerations

### Animal Welfare

The generation and use of knockout mice raises significant animal welfare concerns. Some knockout lines exhibit severe phenotypes, including chronic pain, impaired mobility, or failure to thrive. The principles of the 3Rs—Replacement, Reduction, and Refinement—guide ethical decision-making:

- **Replacement:** Where possible, use alternative models such as cell culture, organoids, or [C. elegans](/knowledge/molecular-biology/c-elegans) or [Yeast Model Organism](/knowledge/molecular-biology/yeast-model-organism) systems that do not involve vertebrate animals.
- **Reduction:** Design experiments to minimize the number of animals used while maintaining statistical power. This includes using appropriate sample sizes and sharing tissues across research groups.
- **Refinement:** Modify procedures to minimize pain and distress. This includes providing appropriate analgesia, environmental enrichment, and humane endpoints.

### Regulatory Guidelines

In the United States, the Animal Welfare Act and the Public Health Service Policy on Humane Care and Use of Laboratory Animals govern the use of mice in research. Institutions must establish an Institutional Animal Care and Use Committee (IACUC) that reviews and approves all animal protocols. In the European Union, Directive 2010/63/EU requires that animal experiments be authorized by competent authorities and that researchers demonstrate the scientific necessity and potential benefits of their work.

## Common Pitfalls and Practical Tips

### Designing Controls

Proper controls are essential for interpreting knockout phenotypes. The ideal control is a wild-type littermate from the same breeding pair, as this controls for maternal effects, litter size, and environmental factors. Heterozygous littermates can also serve as controls, particularly if the knockout allele has a gene-dosage effect. Avoid using mice from different genetic backgrounds or different facilities as controls, as these introduce confounding variables.

### Verifying Knockout

A common pitfall is assuming that the presence of a mutated allele guarantees absence of protein function. Several verification steps are essential:

1. **Confirm the mutation at the DNA level** by sequencing the targeted locus.
2. **Confirm the absence of mRNA** by qRT-PCR using primers spanning the deleted exon.
3. **Confirm the absence of protein** by Western blotting with an antibody recognizing the deleted region.
4. **Confirm the absence of enzymatic activity** if the gene product is an enzyme.

Some knockout strategies produce truncated proteins that retain partial function, and some produce read-through transcripts that bypass the mutation. Without protein-level confirmation, phenotypes may be misinterpreted.

### Interpreting Phenotypes

Phenotypes can be subtle, variable, or absent for reasons unrelated to the gene's true function. Consider the following:

- **Compensation:** Other genes may upregulate to compensate for the loss. This can mask phenotypes or produce phenotypes that reflect compensatory mechanisms rather than the gene's direct function.
- **Environmental factors:** Diet, housing, microbiome, and circadian phase can all influence phenotype. Standardize these factors across groups.
- **Age and sex:** Phenotypes may be age-dependent or sex-specific. Analyze both sexes at multiple time points when feasible.
- **Statistical power:** Knockout phenotypes can be subtle. Use appropriate sample sizes (typically 5–10 animals per group for behavioral studies) and blinded analysis where possible.

## Frequently Asked Questions

### What is a knockout mouse?

A knockout mouse is a genetically engineered mouse in which a specific gene has been inactivated. The gene is disrupted by targeted modification of its sequence, typically by deleting a critical exon, inserting a premature stop codon, or introducing a [frameshift mutation](/knowledge/molecular-biology/frameshift-mutation). This allows researchers to study the gene's function by observing the consequences of its absence.

### How are knockout mice created?

Knockout mice are created by two main methods. The traditional method involves homologous recombination in embryonic stem (ES) cells, where a targeting vector replaces the gene of interest with a selection cassette. Correctly targeted ES cells are injected into blastocysts and implanted into foster mothers, producing chimeric mice that are bred to achieve germline transmission. The modern method uses CRISPR/Cas9, where a guide RNA directs the Cas9 nuclease to introduce a double-strand break at the target locus in fertilized zygotes. The cell's error-prone repair machinery introduces mutations that disrupt the gene.

### What is the difference between a knockout and a knock-in mouse?

A knockout mouse has a gene that has been inactivated or deleted. A knock-in mouse has a gene that has been replaced or modified with a specific variant, such as a point mutation found in human disease, a reporter gene (e.g., GFP), or a human gene sequence. Knock-in mice are used to study the effects of specific mutations rather than complete loss of function.

### Why are knockout mice used in research?

Knockout mice are used to determine gene function, model human diseases, validate drug targets, and study developmental processes. They provide a direct link between genotype and phenotype in a whole organism, which cannot be achieved in cell culture systems. Knockout mice have been instrumental in understanding the genetic basis of diseases such as cancer, diabetes, and neurodegenerative disorders.

### What are conditional knockout mice?

Conditional knockout mice have a gene that is deleted only in specific tissues or at specific times, rather than in all cells throughout development. This is typically achieved using the Cre-loxP system, where the target gene is flanked by loxP sites and Cre recombinase is expressed under a tissue-specific or inducible promoter. Conditional knockouts allow researchers to study genes that are essential for development or to examine gene function in specific cell types.

### Can knockout mice be used to study human diseases?

Yes, knockout mice are widely used to model human genetic diseases. Mice share approximately 85% of their protein-coding genes with humans, and many human disease genes have mouse orthologs. Knockout mice recapitulate many aspects of human diseases, including cystic fibrosis, muscular dystrophy, and various cancers. They are used to understand disease mechanisms and to test potential therapies.

### What are the limitations of knockout mice?

Knockout mice have several limitations. Some gene knockouts are embryonic lethal, preventing analysis in adult animals. Genetic redundancy can mask phenotypes when related genes compensate for the loss. CRISPR/Cas9 can introduce off-target mutations. The genetic background of the mouse strain can influence phenotype. Finally, knockout phenotypes may not fully recapitulate human diseases due to species differences in physiology and gene function.

## Key Takeaways

- A knockout mouse has a specific gene inactivated, allowing direct study of gene function in a whole organism.
- Knockout mice are generated by homologous recombination in embryonic stem cells or by CRISPR/Cas9-mediated genome editing in zygotes.
- Constitutive knockouts delete a gene in all tissues from development, while conditional and inducible knockouts allow tissue-specific or time-controlled deletion.
- Knockout mice are essential for gene function analysis, modeling human diseases, and validating drug targets.
- Phenotypic analysis requires confirmation of knockout at the DNA, RNA, and protein levels, followed by behavioral, molecular, and histological assessments.
- Limitations include embryonic lethality, genetic redundancy, off-target effects, and background strain effects, all of which must be considered in experimental design.
- Ethical use of knockout mice requires adherence to the 3Rs principles and compliance with institutional and regulatory guidelines.

## Further Reading

- van de Wal MAE et al. *Ndufs4 knockout mouse models of Leigh syndrome: pathophysiology and intervention*. Brain : a journal of neurology. 2022. [PubMed 34849584](https://doi.org/10.1093/brain/awab426)
- Lee YJ. *Knockout Mouse Models for Peroxiredoxins*. Antioxidants (Basel, Switzerland). 2020. [PubMed 32098329](https://doi.org/10.3390/antiox9020182)
- Khan S et al. *Inducible Avp knockout mouse line*. American journal of physiology. Renal physiology. 2025. [PubMed 41052028](https://doi.org/10.1152/ajprenal.00340.2025)
- Fienberg AA, Greengard P. *The DARPP-32 knockout mouse*. Brain research. Brain research reviews. 2000. [PubMed 10719158](https://doi.org/10.1016/s0165-0173(99)00047-8)
- Zimmer J, Puri P. *Knockout mouse models of Hirschsprung's disease*. Pediatric surgery international. 2015. [PubMed 26137873](https://doi.org/10.1007/s00383-015-3747-3)
- Mei M, Bao S. *Generation of GM130 Conditional Knockout Mouse*. Methods in [molecular biology](/blog/careers/molecular-biology) (Clifton, N.J.). 2023. [PubMed 36512210](https://doi.org/10.1007/978-1-0716-2639-9_6)

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