STOCK Slc6a3^tm1(cre)Xz/J

Stock number: 020080
Product name: DAT-cre
Research areas: Neurobiology Research, Research Tools

Description

The DAT-cre "knock-in" allele has a nuclear-localized Cre recombinase inserted just upstream of the first coding ATG of the dopamine transporter gene (Slc6a3; DAT). This was designed to both abolish DAT gene function and place Cre recombinase expression under the control of the DAT promoter/enhancer elements.

Detailed description

The DAT-cre "knock-in" allele has a nuclear-localized Cre recombinase inserted just upstream of the translational start codon of the dopamine transporter gene (Slc6a3; DAT). This was designed to both abolish DAT gene function and place Cre recombinase expression under the control of the DAT promoter/enhancer elements.

Progeny derived from crosses with lacZ (see Stock No. 003474) and YFP (see Stock No. 006148) reporter strains confirm that expression is restricted to dopaminergic neurons. Cells in the ventral tegmental area (VTA) and substantia nigra pars compacta (SNc) effectively express Cre and weak expression is also seen in dopaminergic neurons in the olfactory bulb and hypothalamus.

DAT heterozygous knockout mice show subtle behavioral and biochemical changes when compared with wildtype mice. Homozygotes are not healthy and demonstrate a hyperdopaminergic phenotype similar to that of the Slc6a3^tm1Mca knockout (PMID 8628395; Giros, 1996).

The donating investigator notes that unexpected germline Cre expression is occasionally seen in males, but it is rare.

Development

A cassette containing the Cre recombinase coding sequence with a nuclear localization signal and an FRT-flanked neomycin-resistance gene was introduced to the 5' UTR immediately upstream of the translational start codon. The mutation was created via homologous recombination in 129S1/Sv-Oca2⁺Tyr⁺Kitl⁺-derived W9.5 embryonic stem (ES) cells. Resultant chimeric males were crossed with 129Sv/J females. The neomycin cassette was excised through crosses with ACTB:FLP-L mice. This strain was backcrossed to C57BL/6J for more than 10 generations by the donating lab (see SNP note below).

A 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. Two of the 27 markers throughout the genome (on Chromosomes 3 and 12) were segregating, suggesting an incomplete backcross. Also, 3 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.

Allele

Symbol: Slc6a3^tm1(cre)Xz
Name: targeted mutation 1, Xiaoxi Zhuang
MGI accession ID: MGI:3691571
Generation method: Targeted
Attributes: Recombinase-expressing
Marker symbol: Slc6a3
Marker name: solute carrier family 6 (neurotransmitter transporter, dopamine), member 3
Chromosome: 13
Strain of origin: 129S1/Sv-Oca2⁺ Tyr⁺ Kitl⁺
Expressed genes: cre,cre recombinase,bacteriophage P1
Site of expression: Dopaminergic neurons
Molecular note: A targeting vector was constructed by inserting a cassette with the cre recombinase coding sequence, a nuclear translocation signal, and a PGK-neo gene flanked by FRT sites, into the 5'-UTR of the Slc6a3 gene. After homologous recombination in ES cells, mice with cre recombinase expressed from the Slc6a3 gene were produced. Knockin mice were crossed with transgenic Flp deleter mice to remove the neo marker. A line was established where the neo was removed and the Flp transgene was absent. Cre recombinase expression is detected in the ventral tegmental area (VAT) and substantia nigra zona compacta (SNc).