Overview

Also Known As:Luc⁺Lag-3^-

Mice harboring the CAG-luc-eGFP L2G85 transgene and lacking Lag3 exhibit widespread expression of firefly luciferase directed by the CAG promoter, making them useful in studies of transplantation, noninvasive lineage mapping, in vivo bioluminescence imaging and technology development. LAG3 deficiency in these mice enhances the effect of GVHD.

Donating Investigator

Robert Negrin, Stanford University School of Medicine

Genetic overview

Genetic Background	Generation

Lag3^tm1Doi

Allele Type	Gene Symbol	Gene Name
Targeted (Null/Knockout)	Lag3	lymphocyte-activation gene 3

Thy1^a

Allele Type	Gene Symbol	Gene Name
Not Applicable	Thy1	thymus cell antigen 1, theta

Tg(CAG-luc,-GFP)L2G85Chco

Allele Type
Transgenic (Reporter)

Ptprc^a

Allele Type	Gene Symbol	Gene Name
Not Applicable	Ptprc	protein tyrosine phosphatase, receptor type, C

View Genetics

Research Applications

Immunology, Inflammation and Autoimmunity Research
Research Tools

View All Research Applications

Base Price

Starting at:

$2,854.50 Domestic price Cryo Recovery

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Details

Detailed Description

Mice homozygous for the CAG-luc-eGFP L2G85 transgene are viable and fertile, with expression of firefly luciferase and enhanced green fluorescence protein directed by the CAG promoter (human cytomegalovirus immediate early promoter enhancer with chicken beta-actin/rabbit beta-globin hybrid promoter). Exons 1-3 of lymphocyte-activation gene 3 (Lag3) have been replaced with a neomycin resistance gene in reverse orientation to the gene. These mice also harbor congenic Ptprc^a (Cd45.1) and Thy1^a (Thy1.1) cell surface markers allowing for the tracking of these donor cells in vitro. Lag3 gene product is a transmembrane protein expressed in stimulated T lymphocytes and activated natural killer cells. LAG-3 has been found to negatively regulate cellular proliferation, activation, and homeostasis of T cells, and plays a role in Treg suppressive function.

Bioluminesence is detected in heart, spleen, muscle, pancreas, skin, thymus and bone marrow. Luciferase activity is not detected in mature erythrocytes, although low levels are detected in erythrocyte precursors and varying levels of activity in all leukocyte subsets tested. Following luciferin injection, luciferase expression is generally greater in males than females, and the emitted light is adequate to penetrate the tissues of the mice so it can be detected externally with low-light imaging cameras. When mice receive a Lag3 -/- conventional T cell transfer, imaging shows that there is a significant increase in the activation and proliferation compared to transfer of WT conventional T cells. Lag3 deficiency on CD4, but not CD8 T cells, enhances the effect of GVHD. GFP fluorescence is detected in skin (upper epidermal layers) by fluorescence microscopy. The Donating Investigator reports that for the FVB-Tg(CAG-luc,-GFP)L2G85Chco/J (Stock No. 008450) strain, no GFP fluorescence is detected in hematopoietic tissues by flow cytometric analysis.

In an attempt to offer alleles on well-characterized or multiple genetic backgrounds, alleles are frequently moved to a genetic background different from that on which an allele was first characterized. It should be noted that the phenotype could vary from that originally described. We will modify the strain description if necessary as published results become available.

Development

The CAG-luc-eGFP transgenic construct was designed by the laboratory of Dr. Christopher H. Contag (Stanford University School of Medicine). Specifically, the transgene contained the human cytomegalovirus immediate early promoter enhancer with chicken beta-actin/rabbit beta-globin hybrid promoter (CAG; from the pCAGGS vector) upstream of a modified firefly luciferase gene open reading frame, 54 bp of the foot and mouth disease virus (FMDV) 2A sequence (ribsome slippage site), 24 bp polylinker, and enhanced Green Fluorescent Protein (eGFP) open reading frame. This transgene was injected into fertilized FVB mouse eggs. Founder line L2G85 animals were backcrossed to FVB for 20 generations. Dr Robert Negrin obtained these mice and further backcrossed them to B6 for at least 16 generations. These mice also harbor congenic Ptprc^a (Cd45.1) and Thy1^a (Thy1.1) cell surface markers.

Another targeted mutation was developed that replaced exons 1-3 of lymphocyte-activation gene 3 (Lag3) with a neomycin resistance gene in reverse orientation to the gene. The construct was electroporated into 129S2/SvPas-derived D3 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts and the resulting chimeric males were bred to C57BL/6 females. These mice were backcrossed at least five generations to C57BL/6 mice before being bred to the B6 CAG-luc-eGFP mice. Upon arrival at The Jackson Laboratory, mutant mice were bred with C57BL/6J inbred mice (Stock No. 000664) for at least one generation to establish the colony.

Fine mapping performed at The Jackson Laboratory (2011) would indicate that the CAG-luc-eGFP transgene integrated onto the proximal end of chromosome 7 - and the FVB-congenic interval extends from D7Mit306 (~10 Mb) through D7Jmp8 (~29 Mb).

A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. Two of the 27 markers throughout the genome were segregating suggesting an incomplete backcross. One marker on Chr. 7 may be due to transgene insertion. Also, 3 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a C57BL/6 ; C57BL/6N genetic background.

Expression Data

Expressed Gene	GFP, Green Fluorescent Protein,
Expressed Gene	luc, luciferase, firefly
Site of Expression	Widespread expression of firefly luciferase is reported in all tissues and organs, except mature erythrocytes. Following luciferin injection, luciferase expression is generally greater in males than females. GFP fluorescence is detected in skin.
Site of Expression	Widely expressed on all adaptive and innate immune cells.

Control Suggestions

Approximate Controls

000664 C57BL/6J

Additional Information

Considerations for Choosing Controls

Selected References

Sega EI; Leveson-Gower DB; Florek M; Schneidawind D; Luong RH; Negrin RS. 2014. Role of lymphocyte activation gene-3 (Lag-3) in conventional and regulatory T cell function in allogeneic transplantation. PLoS One 9(1):e86551PubMed: 24475140 MGI: J:212715

View All References

Genetics

Lag3^tm1Doi

Allele Symbol: Lag3^tm1Doi

Allele Name	targeted mutation 1, Christophe Benoist and Diane Mathis
Allele Type	Targeted (Null/Knockout)
Allele Synonym(s)	LAG-3^-; Lag3^tm1Dim
Gene Symbol and Name	Lag3, lymphocyte-activation gene 3
Gene Synonym(s)
Site of Expression	Widespread.
Strain of Origin	129S2/SvPas
Chromosome	6
Molecular Note	A neomycin resistance gene replaced a genomic fragment containing exons 1-3. RT-PCR analysis on RNA derived from concanavalin A (Con A) induced splenocytes confirmed that no detectable transcript was produced from this allele.
Mutations Made By	Christophe Benoist, Joslin Diabetes Center

Thy1^a

Allele Symbol: Thy1^a

Allele Name	a variant
Allele Type	Not Applicable
Allele Synonym(s)	thetaAKR; theta-AKR; Thy1.1; Thy-1.1; Thy1a
Gene Symbol and Name	Thy1, thymus cell antigen 1, theta
Gene Synonym(s)
Site of Expression	The Thy1 locus determines a surface antigen present on cells of the thymus, a number of mouse leukemias, brain, and in lesser amounts on lymph node and spleen cells.
Strain of Origin	Not Applicable
Chromosome	9
General Note	The Thy1 locus determines a surface antigen present on cells of the thymus, a number of mouse leukemias, brain, and in lesser amounts on lymph node and spleen cells. The allele Thy1^a determines an antigenic specificity, Thy-1.1, found in the AKR and RF strains; the allele Thy1^b determines an antigenic specificity, Thy-1.2, found in the C3HeB/Fe and many other strains (J:5243, J:5012, J:4469). The Thy1 antigen is probably present on all T lymphocytes and absent from all B lymphocytes, and it thus serves as a valuable T-cell marker (J:5243). It is very widely used in experiments designed to determine the distribution and function of T-cells. Thy1 specifies a T-cell surface glycoprotein, T25, with a molecular weight of 25 kDa (J:5707). The protein appears to be anchored in the cell membrane by a lipid that is either phosphotidylinositol or closely related to it (J:12016). Thy1 may function in the cell membrane as a signal transduction molecule (J:8333). The Thy1 locus, or possibly a gene closely linked to it, controls quantitative expression of a protein that is the same size as Thy1 and is expressed on thymus and brain but not on lymph node and spleen cells (J:7900).
Molecular Note	The allele Thy1^a determines an antigenic specificity, Thy-1.1, found in the AKR and RF strains.

Tg(CAG-luc,-GFP)L2G85Chco

Allele Symbol: Tg(CAG-luc,-GFP)L2G85Chco

Allele Name	transgene insertion L2G85, Christopher H Contag
Allele Type	Transgenic (Reporter)
Allele Synonym(s)	L2685; L2G85; luc⁺; Tg (CAG- Luc, GFP)
Gene Symbol and Name	Tg(CAG-luc,-GFP)L2G85Chco, transgene insertion L2G85, Christopher H Contag
Gene Synonym(s)
Promoter	ACTB, actin, beta, chicken
Promoter	CMV, cytomegalovirus, human
Expressed Gene	GFP, Green Fluorescent Protein,
Expressed Gene	luc, luciferase, firefly
Site of Expression	Widespread expression of firefly luciferase is reported in all tissues and organs, except mature erythrocytes. Following luciferin injection, luciferase expression is generally greater in males than females. GFP fluorescence is detected in skin.
Strain of Origin	FVB
Chromosome	7
General Note	Fine mapping performed at The Jackson Laboratory indicates that the transgene integrated onto the proximal end of Chr. 7 and the congenic interval extends from D7Mit306 (10 Mb) through D7Jmp8 (29 Mb).
Molecular Note	A transgenic construct containing a modified firefly luciferase gene open reading frame, 54bp of the foot and mouth disease virus (FMDV) 2A sequence (ribsome slippage site) and eGFP sequence under the control of the chicken beta actin promoter coupled with the cytomegalovirus (CMV) immediate early enhancer (CAG), was injected into fertilized FVB mouse eggs. Founder line L2G85 animals were bred to wildtype FVB mice.
Mutations Made By	Ryan McBride Spitler, Stanford University

Ptprc^a

Allele Symbol: Ptprc^a

Allele Name	a variant
Allele Type	Not Applicable
Allele Synonym(s)	CD45.1; Ly5^a; Ptprc^SJL
Gene Symbol and Name	Ptprc, protein tyrosine phosphatase, receptor type, C
Gene Synonym(s)
Site of Expression	Widely expressed on all adaptive and innate immune cells.
Strain of Origin	Not Applicable
Chromosome	1
Molecular Note	Ptprc^a is found in strains SJL/J, STS/A, and DA. Ptprc^b is found in strains C57BL/6, C3H/An, DBA/2, AKR, and many others (J:13367, J:12054, J:12077, J:8603). Twelve nucleotide differences between the a and b alleles have been identified. These base substitutions correspond to five amino-acid changes within the extracellular domain of the encoded protein. These amino-acid differences are clustered in a region that also contains the greatest divergence between mouse and rat sequences (J:22485). Note that the allele designations originally described were reversed in 1987 (J:8603); all publications prior to 1987 show SJL/J, STS/A, and DA as having the b allele and the C57BL/6J group as having the a allele (J:22341).

Disease/Phenotype

Disease Terms

Research Areas By Phenotype

This mouse can be used to support research in many areas including:

Genotype: Thy1^a related

Immunology, Inflammation and Autoimmunity Research
- CD Antigens, Antigen Receptors, and Histocompatibility Markers
Research Tools
- Immunology, Inflammation and Autoimmunity Research
  - T cell specific surface marker
- Genetics Research
  - Tissue/Cell Markers: T cell specific surface marker
  - Tissue/Cell Markers

Research Tools
- Fluorescent Proteins
- Cell Biology Research
- Developmental Biology Research
  - transplantation marker for embryonic and adult tissue
- Reproductive Biology Research
  - transplantation marker for embryonic and adult tissue
  - spermatogonial transplantation marker
- Genetics Research
  - Tissue/Cell Markers: cell marker for bone marrow transplantation
  - Tissue/Cell Markers
  - Tissue/Cell Markers: multiple
- Cancer Research
  - xenograft/transplant host
- Toxicology Research
  - B and T cell deficiency, xenograft transplant host
  - xenograft/transplant host

Mammalian Phenotype Terms by Genotype

References

Sega EI; Leveson-Gower DB; Florek M; Schneidawind D; Luong RH; Negrin RS. 2014. Role of lymphocyte activation gene-3 (Lag-3) in conventional and regulatory T cell function in allogeneic transplantation. PLoS One 9(1):e86551PubMed: 24475140 MGI: J:212715

Additional - Lag3^tm1Doi related

Additional - Ptprc^a related

Additional - Thy1^a related

Additional - Tg(CAG-luc,-GFP)L2G85Chco related

Technical Support

CONTACT TECHNICAL SUPPORT

Genotyping Protocols
- Probe:Ptprc End Point-Alternate 1

Breeding Considerations

When maintaining a live colony, these mice can be bred as homozygotes for all four alleles.

Additional Breeding and Husbandry Support

Citation
When using the STOCK Ptprc^a Lag3^tm1Doi Tg(CAG-luc,-GFP)L2G85Chco Thy1^a/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #025855 in your Materials and Methods section.

Animal Health Reports

Facility Barrier Level Descriptions

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Pricing & Availability

Cryo Recovery

Domestic

International

Live Mouse

Age Genotype Price

weeks

Cryorecovery - Pricing

Service/Product Description Price

Homozygous for Ptprc<a> Homozygous for Lag3<tm1Doi>,, Homozygous for Tg(CAG-luc,-GFP)L2G85Chco, Homozygous for Thy1<a>,

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The Jackson Laboratory's Genotype Promise

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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Phone: 207-288-6470

Email: TechTran@jax.org

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Also Known As:Luc+Lag-3-

Donating Investigator

Genetic overview

Research Applications

Base Price

Detailed Description

Development

Expression Data

Control Suggestions

Approximate Controls

Additional Information

Selected References

Lag3tm1Doi

Allele Symbol: Lag3tm1Doi

Thy1a

Allele Symbol: Thy1a

Tg(CAG-luc,-GFP)L2G85Chco

Allele Symbol: Tg(CAG-luc,-GFP)L2G85Chco

Ptprca

Allele Symbol: Ptprca

Disease Terms

Research Areas By Phenotype

This mouse can be used to support research in many areas including:

Genotype: Thy1a related

Mammalian Phenotype Terms by Genotype

References

Additional - Lag3tm1Doi related

Additional - Ptprca related

Additional - Thy1a related

Additional - Tg(CAG-luc,-GFP)L2G85Chco related

Genotyping Protocols

Breeding Considerations

Citation

Animal Health Reports

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Live Mouse

Cryorecovery - Pricing

Related Products and Services

Payment Terms and Conditions

The Jackson Laboratory's Genotype Promise

Terms Of Use

Licensing Information

Also Known As:Luc⁺Lag-3^-

Lag3^tm1Doi

Allele Symbol: Lag3^tm1Doi

Thy1^a

Allele Symbol: Thy1^a

Ptprc^a

Allele Symbol: Ptprc^a

Genotype: Thy1^a related

Additional - Lag3^tm1Doi related

Additional - Ptprc^a related

Additional - Thy1^a related