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| These Ai27D (or Ai27ΔNeo) mice conditionally express an improved channelrhodopsin-2/tdTomato fusion protein (hChR2(H134R)-tdTomato) from the endogenous Gt(ROSA)26Sor locus. Expression is enhanced by the presence of a CAG promoter. Following Cre-mediated removal of the floxed STOP cassette, these mice can be used in optogenetic studies for rapid in vivo activation of excitable cells by illumination with blue light (450-490 nm). | |||||||||||||||||||
- Description
- Disease & phenotype
- Genes & alleles
- Genotyping
- Health & care
- References
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- Terms of use
Description
Strain Information
Type Congenic; Gene Trap; Mutant Strain; Targeted Mutation; Additional information on Genetically Engineered and Mutant Mice. Visit our online Nomenclature tutorial. Additional information on Congenic nomenclature. Mating System Wild-type x Heterozygote (Female x Male) 17-JUL-12 Mating System Heterozygote x Wild-type (Female x Male) 17-JUL-12 Species laboratory mouse Generation N5+N1F1 (15-JUL-11)
Generation DefinitionsDonating Investigator Hongkui Zeng, Allen Institute for Brain Science (AIBS) Description
Ai27D (or Ai27Δneo) mice heterozygous for the Rosa-CAG-LSL-hChR2(H134R)-tdTomato-WPRE conditional allele are viable and fertile. A loxP-flanked STOP cassette prevents transcription of the downstream hChR2(H134R)-tdTomato fusion gene. Because this CAG promoter driven reporter construct was targeted for insertion into the Gt(ROSA)26Sor locus, hChR2(H134R)-tdTomato expression is determined by which tissue(s) express Cre recombinase. When bred to mice that express Cre recombinase, the resulting offspring will have the STOP cassette deleted in the cre-expressing tissues; resulting in expression of the hChR2(H134R)-tdTomato fusion protein. The donating investigator reports that Ai27D mice do not express hChR2(H134R)-tdTomato prior to introduction of Cre recombinase. Fusion protein expression following exposure to cre can be detected by tdTomato fluorescence and mRNA (in situ hybridization) (and presumably by antibody staining (immunohistochemistry); although this was not tested by the donating investigator). Unlike the Ai27 mice from which they were derived, these Ai27D mice no longer harbor the downstream FRT site or attB/attP-flanked selection cassette. The phenotype of homozygous mice has not been characterized by the donating investigator.The hChR2(H134R)-tdTomato fusion protein is composed of a mammalian codon-optimized Chlamydomonas reinhardtii-derived channelrhodopsin-2 that harbors a gain-of-function H134R substitution fused in-frame with a tdTomato fluorescent protein. The hChR2(H134R) is designed to cause larger stationary photocurrents compared to ChR2.
The bacterial opsins are retinal-binding proteins that combine a light-sensitive domain with an ion channel or pump; providing light-dependent ion transport, membrane potential alteration, and sensory functions to bacteria. This hChR2(H134R) functions as a blue light-driven cation channel that depolarizes the cell and causes action potentials. As such, illuminating hChR2(H134R)-expressing cells with blue light (450-490 nm) leads to rapid and reversible photostimulation of action potential firing activity in these cells.
For characterization information, see images at the Allen Institute for Brain Science website (Ai27 images).
Development
The Rosa-CAG-LSL-hChR2(H134R)-tdTomato-WPRE targeting vector was designed with (from 5' to 3') a CMV-IE enhancer/chicken beta-actin/rabbit beta-globin hybrid promoter (CAG), an FRT site, a loxP-flanked STOP cassette (with stop codons in all 3 reading frames and a triple polyA signal), a mammalianized ChR2(H134R)-tdTomato fusion gene, a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE; to enhance the mRNA transcript stability), a BGH polyA signal, and an attB/attP-flanked PGK-FRT-Neo-polyA cassette. To create the mammalianized ChR2(H134R)-tdTomato fusion gene, a cDNA sequence encoding the first 315 amino acids of channelrhodopsin-2 (derived from the green alga Chlamydomonas reinhardtii) was modified to harbor codons optimized for mammalian expression and a gain-of-function H134R substitution (CAC to CGC) designed to cause larger stationary photocurrents. This mhChR2 sequence (also called hChR2(H134R) or hChR2-H134R) was fused in-frame to the amino terminus of a tdTomato sequence (a non-oligomerizing DsRed fluorescent protein variant with a 12 residue linker fusing two copies of the protein (tandem dimer)), resulting in the final hChR2(H134R)-tdTomato fusion protein sequence. The entire Rosa-CAG-LSL-hChR2(H134R)-tdTomato-WPRE targeting vector was inserted between exons 1 and 2 of the Gt(ROSA)26Sor locus via electroporation of (129S6/SvEvTac x C57BL/6)F1-derived G4 embryonic stem (ES) cells. Correctly targeted ES cells (clone Ai27) were selected. Chimeric mice were bred to PhiC31-expressing mice (C57BL/6J congenic background; see Stock No. 007743) to remove the PGK-FRT-Neo-polyA cassette. The resulting Ai27D (or Ai27Δneo) mice were bred with C57BL/6J wildtype mice for at least three more generations (and the PhiC31 transgene was removed) prior to sending to The Jackson Laboratory Repository. Upon arrival, Ai27D mice were bred with C57BL/6J inbred mice (Stock No. 000664) for at least one generations to establish the colony.
Control Information
| Control | ||
|---|---|---|
| Wild-type from the colony | ||
| 000664 C57BL/6J | (approximate) | |
| Considerations for Choosing Controls | ||
Related Strains
Allen Institute for Brain Science cre reporter strains
View Allen Institute for Brain Science cre reporter strains (16 strains)
Additional Web Information
Fluorescent Proteins/lacZ Systems
Introduction to Cre-lox technology
Phenotype
Phenotype Information
assigned by genotype
The following phenotype information may relate to a genetic background differing from this JAX® Mice strain.
Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze/Gt(ROSA)26Sor+
involves: 129S6/SvEvTac * C57BL/6NCr
- no phenotypic analysis
- *normal* no phenotypic analysis (MGI Ref ID J:182204)
This mouse can be used to support research in many areas including:
Neurobiology Research
Cre-lox System
loxP-flanked Sequences
loxP-flanked Sequences: Test/Reporter
Fluorescent protein expression in neural tissue
Optogenetic and Pharmacogenetic tools
Channelrhodopsin expressing strains
Cre-dependent optogenetic tools
Research Tools
Cre-lox System
loxP-flanked Sequences
loxP-flanked Sequences: Test/Reporter
Developmental Biology Research
Cre-lox System
transplantation marker for embryonic and adult tissue
Diabetes and Obesity Research
loxP
Fluorescent Proteins
Genetics Research
Mutagenesis and Transgenesis
Mutagenesis and Transgenesis: Cre-lox System
Tissue/Cell Markers
Tissue/Cell Markers: Cre-lox System
Tissue/Cell Markers: cell marker for bone marrow transplantation
Tissue/Cell Markers: transplantation marker for embryonic and adult tissue
Reproductive Biology Research
Cre-lox System
transplantation marker for embryonic and adult tissue
Genes & Alleles
Gene & Allele Information provided by MGI
| Allele Symbol | Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze | ||
|---|---|---|---|
| Allele Name | targeted mutation 27.1, Hongkui Zeng | ||
| Allele Type | Targeted (Reporter) | ||
| Common Name(s) | Ai27; Ai27D (Rosa-CAG-LSL-hChR2(H134R)-tdTomato-WPRE); Ai27DeltaNeo; ChR2(H134R)-tdTomato; | ||
| Mutation Made By | Hongkui Zeng, Allen Institute for Brain Science (AIBS) | ||
| Strain of Origin | (129S6/SvEvTac x C57BL/6NCr)F1 | ||
| Site of Expression | Cre excision of the stop signal results in expression of an improved channelrhodopsin-2/tdTomato fusion protein (hChR2(H134R)-tdTomato) in cre-expressing tissues. | ||
| Expressed Gene | COP4, Channelrhodopsin, Chlamydomonas | ||
| Molecular Note | The Rosa-CAG-LSL-hChR2(H134R)-tdTomato-WPRE targeting vector was designed with (from 5' to 3') a CMV-IE enhancer/chicken beta-actin/rabbit beta-globin hybrid promoter (CAG), an FRT site, a loxP-flanked STOP cassette (with stop codons in all 3 reading frames and a triple polyA signal), a mammalianized ChR2(H134R)-tdTomato fusion gene, a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE; to enhance the mRNA transcript stability), a BGH polyA signal, and an attB/attP-flanked PGK-FRT-Neo-polyA cassette. To create the mammalianized ChR2(H134R)-tdTomato fusion gene, a cDNA sequence encoding the first 315 amino acids of channelrhodopsin-2 (derived from the green alga Chlamydomonas reinhardtii) was modified to harbor condons optimized for mammalian expression and a gain-of-function H134R substitution (CAC to CGC) designed to cause larger stationary photocurrents. This mhChR2 sequence (also called hChR2(H134R) or hChR2-H134R) was fused in-frame to the amino terminus of a tdTomato sequence (anon-oligomerizing DsRed fluorescent protein variant with a 12 residue linker fusing two copies of the protein (tandem dimer)), resulting in the final hChR2(H134R)-tdTomato fusion protein sequence. The entire Rosa-CAG-LSL-hChR2(H134R)-tdTomato-WPRE targeting vector was inserted between exons 1 and 2 of the locus. PhiC31 mediated recombination removed the PGK-FRT-Neo-polyA cassette. [MGI Ref ID J:182204] | ||
| Gene Symbol and Name | Gt(ROSA)26Sor, gene trap ROSA 26, Philippe Soriano | ||
| Chromosome | 6 | ||
| Gene Common Name(s) | AV258896; Gtrgeo26; Gtrosa26; R26; ROSA26; beta geo; expressed sequence AV258896; gene trap ROSA 26; gene trap ROSA b-geo 26; | ||
Genotyping
Genotyping Information
Genotyping Protocols
Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze,Separated MCA
Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze, Separated PCR
Helpful Links
Genotyping resources and troubleshooting
References
References provided by MGI
Selected Reference(s)
Madisen L; Mao T; Koch H; Zhuo JM; Berenyi A; Fujisawa S; Hsu YW; Garcia AJ 3rd; Gu X; Zanella S; Kidney J; Gu H; Mao Y; Hooks BM; Boyden ES; Buzsaki G; Ramirez JM; Jones AR; Svoboda K; Han X; Turner EE; Zeng H. 2012. A toolbox of Cre-dependent optogenetic transgenic mice for light-induced activation and silencing. Nat Neurosci 15(5):793-802. [PubMed: 22446880] [MGI Ref ID J:182204]
Additional References
Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze relatedPinol RA; Bateman R; Mendelowitz D. 2012. Optogenetic approaches to characterize the long-range synaptic pathways from the hypothalamus to brain stem autonomic nuclei. J Neurosci Methods 210(2):238-46. [PubMed: 22890236] [MGI Ref ID J:192781]
Health & husbandry
Health & Colony Maintenance Information
Animal Health Reports
Room Number AX11
Colony Maintenance
Breeding & Husbandry When maintaining a live colony, heterozygous mice may be bred to wildtype siblings or to C57BL/6J inbred mice (Stock No. 000664). The donating investigator has not tried to generate homozygous mice. Mating System Wild-type x Heterozygote (Female x Male) 17-JUL-12 Heterozygote x Wild-type (Female x Male) 17-JUL-12 Diet Information LabDiet® 5K52/5K67
Pricing and Purchasing
Pricing, Supply Level & Notes, Controls
| Pricing for USA, Canada and Mexico shipping destinations |
|
Live Mice
Price per mouse (US dollars $) Gender Genotypes Provided Individual Mouse $232.00 Female or Male Heterozygous for Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze
Price per Pair (US dollars $) Pair Genotype $302.00 Heterozygous for Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze x Wild-type $302.00 Wild-type x Heterozygous for Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze Standard Supply
Repository-Live.
Repository-Live represents an exclusive set of over 1500 unique mouse models across a vast array of research areas. Breeding colonies provide mice for both large and small orders and fluctuate in size depending on current demand for each strain. If a Repository strain is not immediately available, then within 2 to 3 business days, you will receive an estimated availability timeframe for your inquiry or order along with various delivery options. Repository strains typically are delivered at 4 to 8 weeks of age and will not exceed 12 weeks of age on the day of shipping. We will note and try to accommodate requests for specific ages of Repository strains but cannot guarantee provision of these strains at specific ages. However, if cohorts of mice (5 or more of one gender) are needed at a specific age range for experiments, please let us know.
| Pricing for International shipping destinations |
|
Live Mice
Price per mouse (US dollars $) Gender Genotypes Provided Individual Mouse $301.60 Female or Male Heterozygous for Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze
Price per Pair (US dollars $) Pair Genotype $392.60 Heterozygous for Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze x Wild-type $392.60 Wild-type x Heterozygous for Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hze Standard Supply
Repository-Live.
Repository-Live represents an exclusive set of over 1500 unique mouse models across a vast array of research areas. Breeding colonies provide mice for both large and small orders and fluctuate in size depending on current demand for each strain. If a Repository strain is not immediately available, then within 2 to 3 business days, you will receive an estimated availability timeframe for your inquiry or order along with various delivery options. Repository strains typically are delivered at 4 to 8 weeks of age and will not exceed 12 weeks of age on the day of shipping. We will note and try to accommodate requests for specific ages of Repository strains but cannot guarantee provision of these strains at specific ages. However, if cohorts of mice (5 or more of one gender) are needed at a specific age range for experiments, please let us know.
|
|
|
Standard Supply
Repository-Live.
Repository-Live represents an exclusive set of over 1500 unique mouse models across a vast array of research areas. Breeding colonies provide mice for both large and small orders and fluctuate in size depending on current demand for each strain. If a Repository strain is not immediately available, then within 2 to 3 business days, you will receive an estimated availability timeframe for your inquiry or order along with various delivery options. Repository strains typically are delivered at 4 to 8 weeks of age and will not exceed 12 weeks of age on the day of shipping. We will note and try to accommodate requests for specific ages of Repository strains but cannot guarantee provision of these strains at specific ages. However, if cohorts of mice (5 or more of one gender) are needed at a specific age range for experiments, please let us know.
Control Information
| Control | ||
|---|---|---|
| Wild-type from the colony | ||
| 000664 C57BL/6J | (approximate) | |
| Considerations for Choosing Controls | ||
| Control Pricing Information for Genetically Engineered Mutant Strains. | ||
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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.Ordering Information
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