B6;129S1-Gt(ROSA)26Sor^{tm1(CAG-CD9/GFP)Dmfel}/J

Stock number: 033361
Product name: TIGER , Cre inducible GFP extracellular-vesicle reporter
Strain synonyms: CD9-tGFP^fl
Research areas: Neurobiology Research, Research Tools

Description

TIGER knock-in mice (transgenic inducible GFP EV reporter) express a Cre-inducible His-tagged CD9/TurboGFP reporter targeted to extracellular vesicles.  These mice allow in vivo imaging of extracellular vesicles and are suitable for use in applications related to the study of intercellular communication and transmission of macromolecules between cells.

Detailed description

TIGER mice (transgenic inducible GFP EV reporter) express a Cre-inducible His-tagged CD9/TurboGFP reporter targeted to extracellular vesicles. Exosomes, 50 to 150 nm in size and generated from the endosomal system, and microvesicles, 50 to 500+ nm in size and derived from plasma membrane shedding, are extracellular vesicles. The origin, release, contents, and targeting fate of extracellular vesicles determine the type of cell-cell communication and/or exchange of macromolecules between cells. CD9 is highly expressed in extracellular vesicles.

In the absence of Cre-mediated recombination, neither the His-tagged CD9 nor the GFP reporter is expressed in TIGER mice. Homozygotes are viable and fertile.

Upon crossing to a Cre recombinase-expressing transgene or knock-in, the lox-stop-lox cassette is excised in cells that express cre, leading to production of extracellular vesicle labeling with CD9-GFP. The fluorescent proteins can be visualized based on their intrinsic fluorescence, or by immunostaining for the fluorescent proteins and/or the associated polyhistidine epitope tag.

TurboGFP is an improved variant of the green fluorescent protein CopGFP derived from the copepod Pontellina plumata that exhibits a faster maturation (fluorescence emission) than EGFP. Useful for applications where fast appearance of bright fluorescence is desirable, such as for cell/organelle labeling and tracking promoter activity, TurboGFP has an excitation/emission max of 482/502 nm.

When bred to a strain with tamoxifen-inducible Cre recombinase in adult and developing mouse brain (see Stock No. 016261), this mutant mouse strain may be useful in studies of extracellular vesicles during astrogliogenesis.

Development

A targeting vector containing the CAG promoter, a floxed STOP cassette followed by a carboxy terminal His-tagged CD9/GFP cassette, and a NeoR selection cassette, was used to insert the CAG-LSL-CD9-GFP into the Gt(ROSA)26Sor locus. CD9/GFP is human CD9 containing a carboxy terminal His-tag and fused to copepod green fluorescent protein (CopGFP or Turbo-GFP from Pontellina plumata). The construct was electroporated into B6129S1F1-derived S1B6A embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6J blastocysts. The resulting chimeric animals were bred to C57BL/6J mice, and germline transmission confirmed. The mice were backcrossed to C57BL/6 mice for three generations. During backcrossing, the Y chromosome may not have been fixed to the C57BL/6 genetic background. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).

Allele

Symbol: Gt(ROSA)26Sor^{tm1(CAG-CD9/GFP)Dmfel}
Name: targeted mutation 1, David Feliciano
MGI accession ID: MGI:6392825
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Reporter; Inserted expressed sequence; Epitope tag
Synonyms: TIGER
Marker symbol: Gt(ROSA)26Sor
Marker name: gene trap ROSA 26, Philippe Soriano
Marker synonyms: beta geo; Gtrgeo26; Gtrosa26; R26; ROSA26; SETD5-AS1; Thumpd3as1
Chromosome: 6
Strain of origin: (C57BL/6 x 129S1/SvImJ)F1
Molecular note: A targeting vector containing the CAG promoter, a floxed STOP cassette followed by a carboxy terminal His-tagged CD9/GFP cassette, and a NeoR selection cassette was inserted into the Gt(ROSA)26Sor locus. CD9/GFP is human CD9 containing a carboxy terminal His-tag and fused to copepod green fluorescent protein (CopGFP or Turbo-GFP from Pontellina plumata).
General note: S1B6A embryonic stem cells