B6.Cg-Oxtr^{tm1.1(cre)Hze}/J

Stock number: 031303
Product name: Oxtr-T2A-Cre-D
Research areas: Cancer Research, Neurobiology Research, Research Tools

Description

Oxtr-T2A-Cre-D knock-in mice are designed to have Cre recombinase expression/activity directed by the endogenous oxytocin receptor promoter/enhancer sequences. In the brain, it is observed in restricted populations within the hippocampal formation, cortical subplate, striatum, thalamus, hypothalamus and medulla. Sparse, scattered expression is present in the cortex. These mice may be used to generate conditional mutations for studying gain-or-loss of function and/or fate mapping in OXTR-expressing cells.

Detailed description

The Oxtr gene encodes the oxytocin receptor. Oxtr polymorphisms are of interest in studying human psychology behavioral phenotypes (e.g., autism spectrum disorder).

The Oxtr-T2A-Cre-D knock-in mutation has a T2A sequence and a Cre recombinase gene inserted immediately at the Oxtr translational STOP codon - this is designed to have the endogenous promoter/enhancer elements direct cre expression to OXTR+ cells. When Oxtr-T2A-Cre-D mice are bred with mice containing lox-flanked sequences, Cre-mediated recombination will result in deletion of the floxed sequences in Oxtr-expressing cells in the offspring.

The donating investigator reports Cre recombinase expression/activity largely recapitulates the endogenous Oxtr gene expression in the brain.

Specifically, Cre is enriched in restricted populations within the hippocampal formation, cortical subplate, striatum, thalamus, hypothalamus and medulla. Sparse, scattered expression is present in the cortex. They did not examine cre expression in tissues other than brain.

This knock-in allele is designed to avoid disrupting expression of the endogenous protein; although Oxtr expression from the Oxtr-T2A-Cre-D allele has not been evaluated to date (January 2018).

Heterozygous mice are viable and fertile with no reported gross physical or behavioral abnormalities. To date (January 2018), it has not been attempted to make this strain homozygous.

For characterization information, see images at the Allen Institute for Brain Science website (Oxtr-T2A-Cre-D images).

Development

The Oxtr-T2A-Cre-D mice (also called Oxtr-T2A-Cre-Δ or Oxtr-T2A-Cre-Δneo) were created in the laboratory of Dr. Hongkui Zeng (Allen Institute for Brain Science) to have a T2A sequence and a Cre recombinase gene inserted immediately at the translational STOP codon of the oxytocin receptor gene (Oxtr) on chromosome 6. The specific details are below.

The targeting vector contained, from 5' to 3', a partial Oxtr sequence spanning intron 1 up to but not including the endogenous stop codon in exon 2 (with a frt3 site in intron 1), a viral 2A oligopeptide sequence (T2A; mediates ribosomal skipping), a Cre recombinase gene, a bovine growth hormone polyA sequence, an AttB site, a PGK/gb2 promoter-Neomycin resistance gene-PGK polyA cassette, a frt5 site, an mRNA splice acceptor sequence, the 3' portion of the hygromycin gene (Hygro2) 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. Chimeric mice were bred to PhiC31-expressing mice (C57BL/6J congenic background; see Stock No. 007743) to remove the AttB/AttP-flanked sequences (PGK/gb2-Neo-pA::frt5::RNA splice acceptor::3'hygro-pA) and replace it with the recombined AttB/AttP site (AttL). The resulting Oxtr-T2A-Cre-D mice were bred with C57BL/6J wildtype mice for several generations (and the PhiC31 transgene was removed). In 2018, heterozygous male mice from generation N11 with black coat color were sent to The Jackson Laboratory Repository. Upon arrival, males are used to cryopreserve sperm. To establish the living Oxtr-T2A-Cre-D mouse colony, an aliquot of the frozen sperm is used to fertilize oocytes from C57BL/6J inbred females (Stock No. 000664).
Of note, the donating investigator reports that, at least once during backcrossing, a heterozygous female was bred to a C57BL/6J inbred male (thus the Y chromosome of the congenic strain is of C57BL/6J origin).

Allele

Symbol: Oxtr^{tm1.1(cre)Hze}
Name: targeted mutation 1.1, Hongkui Zeng
MGI accession ID: MGI:6150896
Generation method: Targeted
Attributes: Recombinase-expressing
Synonyms: Oxtr-T2A-Cre-D
Marker symbol: Oxtr
Marker name: oxytocin receptor
Marker synonyms: OTR; OT-R; OTR1
Chromosome: 6
Strain of origin: (129S6/SvEvTac x C57BL/6NCrl)F1
Expressed genes: cre,cre recombinase,bacteriophage P1
Site of expression: When these mice are bred with mice containing lox-flanked sequences, Cre-mediated recombination will result in deletion of the floxed sequences in Oxtr-expressing cells in the offspring.
Molecular note: The targeting vector contained, from 5' to 3', a partial Oxtr sequence spanning intron 1 up to but not including the endogenous stop codon in exon 2 (with a frt3 site in intron 1), a viral 2A oligopeptide sequence (T2A; mediates ribosomal skipping), a Cre recombinase gene, a bovine growth hormone polyA sequence, an AttB site, a PGK/gb2 promoter-Neomycin resistance gene-PGK polyA cassette, a frt5 site, an mRNA splice acceptor sequence, the 3' portion of the hygromycin gene (Hygro2) with SV40 polyA signal, and an AttP site. PhiC31-mediated recombination removed the AttB/AttP-flanked sequences (PGK/gb2-Neo-pA::frt5::RNA splice acceptor::3'hygro-pA) and replace it with the recombined AttB/AttP site (AttL).