B6.Cg-Sst^{tm1.2(tTA2)Hze}/J

Stock number: 016868
Product name: SST-tTA2-D
Research areas: Neurobiology Research, Research Tools

Description

SST-tTA2-D knock-in/knock-out mice have synthetic modified tetracycline-regulated transactivator (tTA2^S) gene expression directed to Sst-expressing cells by the endogenous promoter/enhancer elements of the somatostatin locus. These SST-tTA2-D mice are a Tet-Off tool that allows dox-conditional expression of genes in Sst-expressing cells/tissues.

Detailed description

The SST-tTA2-D (SST-tTA2-Δ) knock-in/knock-out allele both abolishes Sst gene function and expresses a synthetic modified tetracycline-regulated transactivator (tTA2^S) from the somatostatin promoter/enhancer elements.

Heterozygous mice are viable and fertile. Although the donating investigator did not evaluate the phenotype of homozygous animals to date (October 2011), homozygous mice may be expected to have a knockout phenotype similar to other null mutations of this gene and may exhibit nervous system and metabolic abnormalities.

When SST-tTA2-D mice are bred with mice carrying a gene of interest under the regulatory control of a tetracycline-responsive promoter element (TRE, TetRE or tetO), expression of the target gene in the somatostatin-expressing cells may be conditionally abolished with administration of the tetracycline analog doxycycline (dox) in the double mutant offspring. Specifically, the donating investigator reports that SST-tTA2-D activates tetO-regulated gene expression in scattered neural populations, in a pattern that resembles Sst expression. The donating investigator did not examine tTA2^S activity in tissues other than brain.

Development

A targeting vector was used to insert a synthetic modified tetracycline-regulated transactivator (tTA2^S) gene and 3' UTR region of the somatostatin exon 2 into the ATG transcriptional start site of the somatostatin gene (Sst). Specifically, the targeting vector was designed with (from 5' to 3') an frt3 site, a Cre recombinase gene, a bovine growth hormone polyA sequence, a second frt3 site, a tTA2^S gene, the 3' UTR region from somatostatin exon 2, an AttB site, PGK-Neo-polyA cassette, an frt5 site, an RNA splice acceptor, the 3' portion of the hygromycin gene with SV40 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, and chimeric animals were bred to C57BL/6J mice to generate the SST-Cre-pA-tTA2 mutant colony. To remove the frt-flanked Cre recombinase and polyA sequence, the mice were bred to ROSA26::FLPe deleter mice (129S4 genetic background; Stock No. 003946). The SST-tTA2 offspring were then bred to PhiC31-expressing mice (C57BL/6J congenic background; see Stock No. 007743) to replace the PGK-Neo and hygromycin-polyA cassettes with the recombined AttB/AttP site (AttL). The resulting SST-tTA2-D (or SST-tTA2-Δ) mice have a single frt3 site, tTA2^S, the 3' UTR from somatostatin exon 2, and AttL site targeted into the somatostatin ATG site. The SST-tTA2-Δ mice were subsequently backcrossed to C57BL/6J wildtype mice for at least five additional generations (and the FLP-expressing mutation and PhiC31 transgene was removed) prior to sending to The Jackson Laboratory Repository. Upon arrival, mice were bred with C57BL/6J inbred mice (Stock No. 000664) for at least one generation to establish the living colony.

Allele

Symbol: Sst^{tm1.2(tTA2)Hze}
Name: targeted mutation 1.2, Hongkui Zeng
MGI accession ID: MGI:5292254
Generation method: Targeted
Attributes: Transactivator
Marker symbol: Sst
Marker name: somatostatin
Chromosome: 16
Strain of origin: (129S6/SvEvTac x C57BL/6NCrl)F1
Expressed genes: tTA,tetracycline-controlled transactivator,E. coli
Site of expression: Sst^{tm1.2(tTA2)Hze} activates tetO-regulated gene expression in scattered neural populations, in a pattern that resembles Sst expression.
Molecular note: A targeting vector used to insert a synthetic modified tetracycline-regulated transactivator (tTA2S) gene and 3' UTR region of the somatostatin exon 2 into the ATG transcriptional start site of the somatostatin gene (Sst). Specifically, the targeting vector was designed with (from 5' to 3') an frt3 site, a Cre recombinase gene, a bovine growth hormone polyA sequence, a second frt3 site, a tTA2S gene, the 3' UTR region from somatostatin exon 2, an AttB site, PGK-Neo-polyA cassette, an frt5 site, an RNA splice acceptor, the 3' portion of the hygromycin gene with SV40 polyA signal, and an AttP site.