B6;129S-Slc17a7^{tm1.1(cre)Hze}/J

Stock number: 023527
Product name: Slc17a7-IRES2-Cre-D or Vglut1-IRES2-Cre-D
Research areas: Cell Biology Research, Neurobiology Research, Research Tools

Description

Slc17a7-IRES2-Cre-D knockin mice have Cre recombinase expression directed to Vglut1-expressing cells, without disrupting endogenous vesicular glutamate transporter 1 expression. These mice may be useful for studying glutamatergic synaptic vesicle trafficking and vesicle-bound, sodium-dependent phosphate transportation.

Detailed description

The Slc17a7-IRES2-Cre-D (also called Slc17a7-IRES2-Cre-Δ or Slc17a7-IRES2-Cre-Δneo/hygro) targeted mutation has an IRES2 sequence and a Cre recombinase gene inserted downstream of the translational STOP codon of the vesicular glutamate transporter 1 gene (Slc17a7 or Vglut1). As such, Slc17a7-IRES2-Cre-D mice have both endogenous gene and Cre recombinase expression directed to Slc17a7-expressing cells by the endogenous promoter/enhancer elements of the vesicular glutamate transporter 1 locus. When Slc17a7-IRES2-Cre-D mice are bred with mice containing loxP-flanked sequences, Cre-mediated recombination will result in deletion of the floxed sequences in Slc17a7-expressing cells in the offspring.

The donating investigator reports Cre recombinase expression (in situ hybridization using a Cre-specific probe) is observed in a pattern very similar to that of the endogenous Slc17a7 gene. Specifically, cre expression is strong throughout the cortex, olfactory bulb and anterior olfactory nuclei. Scattered cre expression is seen in striatum and hippocampus. Cre recombinase is enriched in restricted populations in pons, superior colliculus and anterodorsal nucleus of thalamus. The donating investigator did not examine cre expression in tissues other than brain. Heterozygous mice are viable and fertile with no reported gross physical or behavioral abnormalities. The donating investigator has not attempted to generate homozygous mice to date (September 2013).

For characterization information, see images at the Allen Institute for Brain Science website (Slc17a7-IRES2-Cre images).

Development

These Slc17a7-IRES2-Cre-D mice (also called Slc17a7-IRES2-Cre-Δ or Slc17a7-IRES2-Cre-Δneo/hygro) have an IRES2 sequence and a Cre recombinase gene inserted immediately downstream of the translational STOP codon of the vesicular glutamate transporter 1 gene (Slc17a7 or Vglut1). The specific details are below.

The targeting vector contained, from 5' to 3', a partial Slc17a7 sequence spanning exon 1 through intron 9, an frt3 site within Slc17a7 intron 9, a partial Slc17a7 exon 10 sequence up to and including the endogenous stop codon, an internal ribosome entry site 2 (IRES2) sequence (allows translation initiation in the middle of an mRNA sequence), a Cre recombinase gene, a bovine growth hormone polyA sequence, an AttB site, a PGK promoter-Neomycin resistance gene-PGK polyA cassette, an frt5 site, an mRNA splice acceptor, the 3' portion of the hygromycin gene with SV40 late polyA signal, and an AttP site.

This construct was electroporated into (129S6/SvEvTac x C57BL/6)F1-derived G4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts. Chimeric mice were bred to PhiC31-expressing mice (C57BL/6J congenic background; see Stock No. 007743) to remove the AttB/AttP-flanked sequences (PGK-Neo-polyA::frt5::RNA splice acceptor::3'hygro-polyA) and replace it with the recombined AttB/AttP site (AttL).

The resulting Slc17a7-IRES2-Cre-D mice were bred with C57BL/6J wildtype mice for several generations (and the PhiC31 transgene was removed). In 2013, males from generation N3F1 were sent to The Jackson Laboratory Repository. Upon arrival, males were used to cryopreserve sperm. To establish the living Slc17a7-IRES2-Cre-D mouse colony, an aliquot of the frozen sperm was used to fertilize oocytes from C57BL/6J inbred females (Stock No. 000664).

Allele

Symbol: Slc17a7^{tm1.1(cre)Hze}
Name: targeted mutation 1.1, Hongkui Zeng
MGI accession ID: MGI:5507862
Generation method: Targeted
Attributes: Recombinase-expressing
Marker symbol: Slc17a7
Marker name: solute carrier family 17 (sodium-dependent inorganic phosphate cotransporter), member 7
Chromosome: 7
Strain of origin: (129S6/SvEvTac x C57BL/6NCrl)F1
Expressed genes: cre,cre recombinase,bacteriophage P1
Site of expression: cre expression is strong throughout the cortex, olfactory bulb and anterior olfactory nuclei. Scattered cre expression is seen in striatum and hippocampus. Cre recombinase is enriched in restricted populations in pons, superior colliculus and anterodorsal nucleus of thalamus.
Molecular note: The targeting vector contained, from 5' to 3', a partial Slc17a7 sequence spanning exon 1 through intron 9, an frt3 site within Slc17a7 intron 9, a partial Slc17a7 exon 10 sequence including the endogenous stop codon, an internal ribosome entry site 2 (IRES2) sequence , a Cre recombinase gene, a bovine growth hormone polyA sequence, an AttB site, a PGK promoter-Neomycin resistance gene-PGK polyA cassette, an frt5 site, an mRNA splice acceptor, the 3' portion of the hygromycin gene with SV40 late polyA signal, and an AttP site. This construct was electroporated into G4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts. Chimeric mice were bred to PhiC31-expressing mice to remove the AttB/AttP-flanked sequences (PGK-Neo-polyA::frt5::RNA splice acceptor::3'hygro-polyA) and replace it with the recombined AttB/AttP site (AttL). The resulting Slc17a7-IRES2-Cre-D mice were bred with C57BL/6J wildtype mice for several generations to remove PhiC31 transgene.