This triple mutant strain carries a humanized ApoE knock-in mutation (sequence coding for isoform E4), a CRISPR/Cas9-generated 387bp deletion in the first coding exon knock-out mutation of the Il1rap gene and a CRISPR/cas9-generated R47H point mutation of the Trem2 gene. These mice may be suitable for use in studies related to Alzheimer's disease, lipoproteins, arteriosclerosis, and coronary heart disease.
Mike Sasner, The Jackson Laboratory
Genetic Background | Generation |
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|
Allele Type | Gene Symbol | Gene Name |
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Endonuclease-mediated (Humanized sequence) | Trem2 | triggering receptor expressed on myeloid cells 2 |
Allele Type | Gene Symbol | Gene Name |
---|---|---|
Targeted (Inserted expressed sequence, Humanized sequence) | Apoe | apolipoprotein E |
Allele Type | Gene Symbol | Gene Name |
---|---|---|
Endonuclease-mediated (Null/Knockout) | Il1rap | interleukin 1 receptor accessory protein |
This triple mutant strain carries a humanized ApoE knock-in allele, in which exons 2, 3 and most of exon 4 of the mouse Apoe gene were replaced by human APOE4 gene sequence including exons 2, 3 and 4 (and some 3' UTR sequence); a mutant knock-out allele, 387bp deletion in the first coding exon, of the Il1rap gene; and a knock-in of a point mutation into mouse Trem2, triggering receptor expressed on myeloid cells 2, gene containing a R47H point mutation, with two silent mutations. The targeted Apoe gene encodes apolipoprotein E, which is important in lipoprotein metabolism and cardiovascular disease as well as Alzheimer's disease, immunoregulation and cognition. The targeted Il1rap gene encodes interleukin 1 receptor accessory protein which is a component of the interleukin 1 receptor complex, and is involved in interleukin 1 signalling. The targeted Trem2 gene encodes a protein that is part of a receptor signaling complex with TYRO protein tyrosine kinase binding protein, and that activates macrophages and dendritic cells during immune responses. The TREM2 R47H mutation is a missense mutation in exon 2 that is one of the strongest genetic risk factors for late-onset Alzheimer's disease. Mice that are homozygous for the Apoetm1.1(APOE*4)Adiuj and Trem2em1Adiuj alleles, and heterozygous for the Il1rapem1Adiuj allele are viable and fertile. Homozygous viability/fertility has not been tested for the Il1rapem1Adiuj allele (June 2018). As the mice are characterized, we will modify the strain description and add phenotype data.
Of note, in brains of mice homozygous for the Trem2em1Adiuj allele (and not carrying any other mutant alleles), expression of both transcripts of Trem2 is decreased by about 50%. Mice expressing the Trem2 R47H mutation also express a novel splice variant with a deletion of 119bp at the 5’ end of exon 2, due to a cryptic splice acceptor site in exon 2 (see Stock No. 027918).
This triple mutant line was generated by utilizing CRISPR/cas9 endonuclease mediated genome editing of the Il1rap gene in B6(SJL)-Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J (available as Stock No. 028709) mice.
The double mutant B6(SJL)-Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J (Stock No. 028709) was generated by crossing 2 single mutant lines.
For the Apoetm1.1(APOE*4)Adiuj allele (available as Stock No. 027894):
A targeting vector containing a 3' FRT flanked neo cassette was used to replace exons 2, 3 and most of exon 4 with 1.5 kb of human APOE gene sequence including exons 2, 3 and 4 (and some 3' UTR sequence). The human ApoE sequence also contains a 2 nucleotide A to T substitution in intron 3 for identification of the targeted allele by Southern analysis. The construct was electroporated into C57BL/6J derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric animals were tested for germline transmission by crossing to C57BL/6J. Heterozygous animals were crossed subsequently to FLP recombinase expressing mice (see Stock No. 005703) to remove the FRT site flanked PGK-neo cassette. Mice that no longer contained the FRT flanked PGK-neo cassette were then backcrossed to C57BL/6J at least once to remove the FLP recombinase transgene.
For the Trem2em1Adiuj allele (available as Stock No. 027918):
Plasmids encoding a single guide RNA designed to introduce a R47H point mutation, with two silent mutations, into the Trem2 gene and the cas9 nuclease were introduced into the cytoplasm of C57BL/6J-derived fertilized eggs with well recognized pronuclei. Correctly targeted embryos were transferred to pseudopregnant females. Correctly targeted pups were identified by sequencing and PCR and further bred to C57BL/6J (Stock No. 000664) to develop the colony.
For the Il1rapem1Adiuj allele: plasmids encoding a single guide RNA designed to introduce a knock-out mutation, 387bp deletion in the first coding exon, into the Il1rap gene and the cas9 nuclease were introduced into the cytoplasm of B6(SJL)-Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J (Stock No. 028709)-derived fertilized eggs with well recognized pronuclei. Correctly targeted embryos were transferred to pseudopregnant females. Correctly targeted pups were identified by sequencing and PCR and further bred to B6(SJL)-Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J (Stock No. 028709) to develop the colony.
Expressed Gene | APOE, apolipoprotein E, human |
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Site of Expression | Endogenous mouse elements direct human APOE gene sequence (including exons 2, 3 and 4 and some 3' UTR sequence). |
Site of Expression |
Allele Name | endonuclease-mediated mutation 1, MODEL-AD Center |
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Allele Type | Endonuclease-mediated (Humanized sequence) |
Allele Synonym(s) | Jax R27H ki; Trem2 R47H KI |
Gene Symbol and Name | Trem2, triggering receptor expressed on myeloid cells 2 |
Gene Synonym(s) | |
Strain of Origin | C57BL/6J |
Chromosome | 17 |
Molecular Note | The allele was generated by injecting cas9 nuclease and a single guide RNA designed to introduce a R47H point mutation with two silent mutations (lysine AAG>AAA and alanine GCC>GCA) into the gene. |
Allele Name | targeted mutation 1.1, MODEL-AD Center |
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Allele Type | Targeted (Inserted expressed sequence, Humanized sequence) |
Allele Synonym(s) | APOEepsilon4 |
Gene Symbol and Name | Apoe, apolipoprotein E |
Gene Synonym(s) | |
Expressed Gene | APOE, apolipoprotein E, human |
Site of Expression | Endogenous mouse elements direct human APOE gene sequence (including exons 2, 3 and 4 and some 3' UTR sequence). |
Strain of Origin | C57BL/6J |
Chromosome | 7 |
Molecular Note | The targeting vector contains a 3' FRT flanked neo cassette and 1.5 kb of human APOE gene sequence including exons 2, 3, 4 and a portion of the 3'UTR sequence (the APOE4 isoform) which is used to replace exons 2, 3 and most of exon 4 of the endogenous sequence. The human ApoE sequence carries a 2 nucleotide A to T substitution in intron 3. Flp-mediated recombination removed the FRT-flanked neo cassette. |
Allele Name | endonuclease-mediated mutation 1, MODEL-AD Center |
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Allele Type | Endonuclease-mediated (Null/Knockout) |
Allele Synonym(s) | |
Gene Symbol and Name | Il1rap, interleukin 1 receptor accessory protein |
Gene Synonym(s) | |
Strain of Origin | B6(SJL)-Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J |
Chromosome | 16 |
Molecular Note | CRISPR/cas9 endonuclease-mediated genome editing is used to introduce a 387bp deletion in the first coding exon. The mutation is believed to generate a null allele. |
When maintaining a live colony, mice homozygous for the Apoetm1.1(APOE*4)Adiuj and Trem2em1Adiuj alleles, and heterozygous for the Il1rapem1Adiuj allele may be bred.
Homozygous viability/fertility has not been tested for the Il1rapem1Adiuj allele (June 2018).
As the mice are characterized, we will modify the strain description if necessary and add data related to viability and fertility.
When using the Il1rap KO/APOE4/Trem2*R47H, exon 2 KO
mouse strain in a publication, please cite the originating article(s) and include JAX stock #030304 in your Materials and Methods section.
Facility Barrier Level Descriptions
Service/Product | Description | Price |
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Homozygous for Apoe<tm1.1(APOE*4)Adiuj> and Trem2<em1Adiuj>, heterozygous or wildtype for Il1rap<em1Adiuj> |
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The Jackson Laboratory has rigorous genetic quality control and mutant gene genotyping programs to ensure the genetic background of JAX® Mice strains as well as the genotypes of strains with identified molecular mutations. JAX® Mice strains are only made available to researchers after meeting our standards. However, the phenotype of each strain may not be fully characterized and/or captured in the strain data sheets. Therefore, we cannot guarantee a strain's phenotype will meet all expectations. To ensure that JAX® Mice will meet the needs of individual research projects or when requesting a strain that is new to your research, we suggest ordering and performing tests on a small number of mice to determine suitability for your particular project. We do not guarantee breeding performance and therefore suggest that investigators order more than one breeding pair to avoid delays in their research.
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