B6.Cg-Slc6a9^{em1(icre)Mira}/J

Stock number: 040344
Product name: GlyT1-CRE
Research areas: Cell Biology Research, Neurobiology Research, Research Tools

Description

GlyT1-CRE-iCre knock-in mice have a P2A sequence and codon-optimized cre recombinase gene inserted into exon 14 of the Slc6a9 gene, while retaining endogenous Slc6a9 expression. These mice may be useful for tracking Slc6a9-expressing cells of the central nervous system.

Detailed description

GlyT1-CRE-iCre knock-in mice have a P2A sequence and codon-optimized cre recombinase gene inserted into exon 14 of the solute carrier family 6 (neurotransmitter transporter, glycine), member 9 (Slc6a9) gene, while retaining endogenous Slc6a9 expression. Glycine transporter 1 (GlyT1), coded by Slc6a9, is a sodium/chloride-dependent transporter responsible for regulating levels of the inhibitory neurotransmitter, glycine, as well as playing a role in N-methyl-D-aspartate (NMDA) receptor function.

Homozygous mice are viable and fertile, with no overt phenotype. When these GlyT1-CRE-iCre mice are bred with mice containing loxP-flanked sequence, Cre-mediated recombination will result in deletion of the floxed sequences in the cre-expressing cells of the offspring.

The donating laboratory bred these GlyT1-CRE-iCre knock-in mice to the Ai14 tdTomato reporter line (Stock No. 007914) or to the Ai6 ZsGreen reporter line (Stock No. 007906) to evaluate Cre activity. Resulting progeny demonstrated fluorescence throughout areas of the brain stem, cerebellum, the cerebral peduncles and the forebrain such as the thalamus, striatum, cortex and the basal ganglia regions, correlating with endogenous Slc6a9 expression. The donating investigator also notes Cre activity in the pancreas, skin and retina. These mice may be useful for tracking Slc6a9-expressing cells.

Development

The GlyT1-CRE knock-in allele was generated using CRISPR/Cas9 endonuclease-mediated genome editing. Guide RNAs were used to introduce a P2A peptide, followed by codon-optimized iCre-recombinase and polyA sequences, into exon 14 of the solute carrier family 6 (neurotransmitter transporter, glycine), member 9 (Slc6a9) gene on chromosome 4 (MGI:95760). The sgRNAs, ssDNA and the Cas9 nuclease were introduced into the cytoplasm of zygotes derived from B6D2F1/J (Stock No. 100006) mice with well recognized pronuclei. Correctly targeted pups were identified by PCR and sequencing. The resulting GlyT1-CRE knock-in mice were bred to C57BL/6J (Stock No. 000664) mice for 10 generations before being crossed to either the Ai14 tdTomato reporter line (Stock No. 007914) or to the Ai6 ZsGreen reporter line (Stock No. 007906), by the donating laboratory. The donor mice sent to The Jackson Laboratory, were heterozygous for GlyT1-CRE and wildtype for Ai14 tdTomato and Ai6 ZsGreen. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes.

Allele

Symbol: Slc6a9^{em1(icre)Mira}
Name: endonuclease-mediated mutation 1, Manuel Miranda
MGI accession ID: MGI:8246644
Generation method: Endonuclease-mediated
Attributes: Recombinase-expressing
Marker symbol: Slc6a9
Marker name: solute carrier family 6 (neurotransmitter transporter, glycine), member 9
Marker synonyms: GCENSG; GLYT1; Glyt-1; GLYT-1b; IS6
Chromosome: 4
Strain of origin: (C57BL/6J x DBA/2J)F1/J
Molecular note: Using CRISPR/Cas9 endonuclease-mediated genome editing, guide RNAs were used to introduce a P2A peptide, followed by codon-optimized iCre-recombinase and polyA sequences, into exon 14 of the solute carrier family 6 (neurotransmitter transporter, glycine), member 9 (Slc6a9) gene.