B6.Cg-Tg(Gfap-flpo)62Thara/J

Stock number: 033116
Product name: GFAP-FLPo line #62
Research areas: Developmental Biology Research, Neurobiology Research, Research Tools

Description

GFAP-FLPo transgenic mice have the mouse glial fibrillary acidic protein gene sequences directing expression of optimized FLP recombinase (FLPo) in specific GFAP+ populations such as astrocytes and, to a lesser extent, neural stem cells (NSCs). These GFAP-FLPo line #62 mice allow in vivo genetic manipulations for studying brain biology at different developmental stages in neural circuitry/synapses, behavior, injury/trauma and disease (e.g., gliomas).

Detailed description

Glial fibrillary acidic protein (GFAP) is a cell-specific marker expressed in astrocytes and neural stem cells (NSCs). GFAP+ astrocytes and NSCs have been shown to be cell-of-origins of gliomas in the adult mouse brain.

The GFAP-FLPo transgene has the mouse glial fibrillary acidic protein gene sequences directing expression of an optimized FLP recombinase (FLPo). GFAP-FLPo mice from founder line #62 exhibit FLPo recombinase expression/activity in specific GFAP+ populations such as astrocytes and, to a lesser extent, neural stem cells (NSCs). Specific details are described below.
When GFAP-FLPo line #62 are bred with mice containing frt-flanked sequences, FLPo-mediated recombination will result in deletion of the frt-flanked sequences in Gfap-expressing cells in the offspring. The donating investigator reports FLPo recombinase expression/activity largely recapitulates the endogenous Gfap gene in the brain in terms of tissue specificity, however some GFAP+ cells can be negative for FLPo expression based on lineage tracing experiments. While GFAP-FLPo line #62 was originally found to have the least amount of non-brain (spleen) expression of all founder lines examined, the donating investigator did not examine FLPo expression in tissues other than brain (such as central nervous system or germline). Of note, other mouse lines using the same Gfap promoter exhibit male germline expression - see Stock Nos. 012886/024098.

When GFAP-FLPo line #62 are bred to reporter mice that have FLP-dependent tdTomato expression (RC::FLTG; Stock No. 026932), the resulting offspring demonstrate tightly-controlled FLPo expression in specific GFAP+ populations such as astrocytes. Overall, FLPo expression/activity was significantly biased toward GFAP+ astrocytes in the hippocampus, although limited FLPo-mediated recombination was observed in NSCs in the subgranular zone (SGZ) of the hippocampal dendate gyrus.

Hemizygous GFAP-FLPo line #62 mice are viable and fertile with normal breeding, and no reported phenotypic abnormalities. To date (April 2019), it has not been attempted to make this strain homozygous.

Development

GFAP-FLPo transgenic mice were designed in the laboratory of Dr. Toshiro Hara (Salk Institute). First, a 15 kbp mouse glial fibrillary acidic protein (Gfap) sequence was obtained including its promoter region, regulatory elements, exons, introns and 3' UTR, as well as having a point mutation (ATG to TTG) in the Gfap start codon site to prevent GFAP expression from the transgene. To this sequence, an optimized FLP recombinase (FLPo) followed by BGH polyA was integrated into the Gfap first exon. The final ~14.7 kbp GFAP-FLPo-bGHpoly(A) transgene was used for pronuclear injection into B6D2F1/J x B6D2F1/J embryos. Founder animals were bred to C57BL/6J inbred mice for germline transmission. Founder line #62 was identified to have FLPo expression predominantly in brain, and had the least amount of non-brain expression. The donating investigator reports that GFAP-FLPo line #62 was backcrossed at least nine generations to C57BL/6J inbred mice, and then hemizygous males with black coat color were sent to The Jackson Laboratory Repository in 2019. Upon arrival, males are used to cryopreserve sperm. To establish our living colony, an aliquot of the frozen sperm is used to fertilize oocytes from C57BL/6J inbred females (Stock No. 000664).
Of note, it is not known if the Y chromosome has been fixed to the C57BL/6J background during backcrossing. The transgene insertion site(s) and copy number are not known to date (April 2019).

Allele

Symbol: Tg(Gfap-flpo)62Thara
Name: transgene insertion 62, Toshiro Hara
MGI accession ID: MGI:6294098
Generation method: Transgenic
Attributes: Recombinase-expressing
Synonyms: GFAP-FLPo
Marker symbol: Tg(Gfap-flpo)62Thara
Marker name: transgene insertion 62, Toshiro Hara
Chromosome: UN
Strain of origin: B6D2F1
Molecular note: A transgene consist of an 15 kbp mouse glial fibrillary acidic protein (Gfap) sequence including its promoter region, regulatory elements, exons, introns and 3' UTR, as well as having a point mutation (ATG to TTG) in the Gfap start codon site to prevent GFAP expression from the transgene with an optimized FLP recombinase (flpo) followed by BGH polyA integrated into the Gfap first exon was used. The final transgene was used for pronuclear injection into B6D2F1/J x B6D2F1/J embryos. Founder animals were bred to C57BL/6J inbred mice for germline transmission. Founder line #62 was identified to have flpo expression predominantly in brain, and had the least amount of non-brain expression.