B6.Cg-Kcnma1^tm1.1Alme/J

Stock number: 035901
Product name: Kcnma1^fl-tdTomato
Research areas: Cell Biology Research, Neurobiology Research, Research Tools

Description

Kcnma1^fl-tdTomato conditional mutant mice possess loxP sites flanking exon 2 and a tdTomato reporter (that only expresses upon cre recombination) in the Kcnma1 (potassium large conductance calcium-activated channel, subfamily M, alpha member 1) gene and may be useful in generating conditional mutations to study the function of BK currents.

Detailed description

Kcnma1 (potassium large conductance calcium-activated channel, subfamily M, alpha member 1) encodes the pore-forming alpha subunit of the BK voltage and calcium-sensitive potassium channel (K_Ca1.1). BK channels regulate membrane potential and repolarization of action potentials and are widely expressed in neurons,smooth muscle, neuroendocrine, and non-excitable cells such as bone and kidney. Mutations in Kcnma1 are associated with paroxysmal non-kinesigenic dyskinesia (PKND3) and epileptic seizures, as well as neurodevelopmental disorder.

These conditional Kcnma1^fl-tdTomato mice possess loxP sites flanking exon 2 and a 2A-tdTomato reporter in reverse orientation. Mice that are homozygous for this floxed allele are viable and fertile. When bred to mice that express tissue-specific Cre recombinase, resulting offspring will have exon 2 inverted and the 2A-tdTomato reporter brought in frame (and expressed) in cre-expressing tissues.
For example, when bred to mice with Cre recombinase expression in neurons of the central and peripheral nervous systems (for example, Tg(Nes-cre)1Kln and SM222a-Cre), homozygous knockout offspring were verified to lack BK currents in neurons of the suprachiasmatic nucleus and mesenteric artery smooth muscle cells.

Development

The targeting construct was designed to insert a lox71 site upstream of exon 2, and an FRT-flanked neomycin resistance cassette in reverse orientation followed by 2A-tdTomato-polyA sequence also in reverse orientation and a lox66 site into the Kcnma1 gene on chromosome 14. The construct was electroporated into hybrid (C57BL/6 x 129S/SvEv)F1-derived BA1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric males were crossed to B6.SJL-Tg(ACTFLPe)9205Dym/J to remove the neomycin cassette. Progeny were backcrossed to C57BL/6J for 11 generations and the FLP allele bred out. Upon arrival, mice were bred to C57BL/6J for at least 1 generation to establish the colony.

Allele

Symbol: Kcnma1^tm1.1Alme
Name: targeted mutation 1.1, Andrea L Meredith
MGI accession ID: MGI:5911897
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Reporter; No functional change
Marker symbol: Kcnma1
Marker name: potassium large conductance calcium-activated channel, subfamily M, alpha member 1
Chromosome: 14
Strain of origin: (C57BL/6NTac x 129S6/SvEvTac)F1
Molecular note: A lox71 site was inserted upstream of exon 2 and an FRT flanked neo cassette, 2A-tdTomato gene and lox66 site in inverse orientation were inserted downstream of exon 2. Flp mediated recombination removed the neo cassette. In the absence of cre-mediated recombination the tdTomato reporter is not expressed. Following cre-mediated the tdTomato reporter is expressed and exon 2 is placed in inverse orientation. The cre-mediated inversion is irreversible.