B6.Cg-Pvalb^tm1(cre)Arbr Fxn^em2Lutzy Fxn^em2.1Lutzy/J

Stock number: 029721
Product name: Fxn^flox/null::PV-Cre
Research areas: Neurobiology Research, Research Tools

Description

Fxn^flox/null::PV-Cre mice have a Cre-conditional frataxin allele, global knockout frataxin allele and parvalbumin neuron-specific Cre recombinase knockin allele - useful to study Friedreich's Ataxia.

The Fxn^flox/null::PV-Cre genotype is compound heterozygous at the frataxin locus (floxed exon 2 and global KO on respective homologous chromosomes) and heterozygous for the PV-Cre knockin allele. Due to early-onset ataxia, Fxn^flox/null::PV-Cre are distributed at 4-7 weeks of age.
In addition, we distribute the phenotypically-normal parental lines:
i. Stock No. 028520: C57BL/6J-congenic Fxn^em2Lutzy (homozygous for floxed exon 2)
ii. Stock No. 030218: C57BL/6-congenic Pvalb^tm1(cre)Arbr Fxn^em2.1Lutzy (heterozygous for both PV-Cre knockin and frataxin global KO).
These breeders produce the affected Fxn^flox/null::PV-Cre genotype at a 1/4 frequency.

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Detailed description

The Fxn gene encodes mitochondrial frataxin protein which is involved in mitochondrial iron transport and respiration regulation. Mutations in this gene have been associated with Friedreich's Ataxia. These mice carry the Pvalb^tm1(cre)Arbr allele, expressing Cre recombinase in parvalbumin-expressing neurons, the Fxn^flox (floxed exon 2; Fxn^em2Lutzy) allele and the Fxn^null (global Fxn knock-out; Fxn^em2.1Lutzy) allele. This triple mutant strain is designed to generate mice that are heterozygous for the Fxn^null (global knock-out Fxn) allele with ablation of Fxn expression in parvalbumin-expressing neurons. These mice exhibit an observable ataxic phenotype onset at approximately 9 to 10 weeks of age and have a mean survival of approximately 17 weeks.

Of note, parvalbumin is expressed in sperm. However, characterization of Pvalb^tm1(cre)Arbr mice performed by The Jackson Laboratory Genetic Resource Science group reports no Cre expression in spermatids, and Pvalb^tm1(cre)Arbr did not exhibit germline recombination in multiple crosses from different labs - see Stock No. 029721 Detailed Description for more information.

Development

This triple mutant line was generated by breeding female mice carrying the Pvalb^tm1(cre)Arbr (see Stock No. 017320) and Fxn^em2.1Lutzy (see Stock No. 028040) alleles to male C57BL/6J-Fxn^em2Lutzy/J mice (Stock No. 028520). (As parvalbumin is expressed in sperm there is ectopic Cre recombinase expression in the male germline of mice carrying the Pvalb^tm1(cre)Arbr allele.)

The frataxin floxed exon 2 allele Fxn^flox (Fxn^em2Lutzy) was made by Dr. Cat Lutz at The Jackson Laboratory using CRISPR/cas9 endonuclease-mediated genome editing. Plasmids encoding a signal guide RNA designed to introduce loxP sites flanking exon 2 in the Fxn gene and the CRISPR/cas9 nuclease were introduced into the cytoplasm C57BL/6J-derived fertilized eggs with well recognized pronuclei. Correctly targeted embryos were transferred to pseudopregnant females. Correctly targeted pups were identified by sequencing and PCR. Mice from founder 466 were further bred to C57BL/6J inbred mice (Stock No. 000664) to develop the frataxin floxed exon 2 colony as Stock No. 028520.

To generate the global knockout allele Fxn^null (Fxn^em2.1Lutzy), male mice carrying the floxed exon 2 allele (from Stock No. 028520) were bred to female B6N.Cg-Tg(Sox2-cre)1Amc/J (Stock No. 014094). The resulting mice (exon 2 deleted from germline) were further bred to C57BL/6J to breed out the Cre recombinase transgene. Heterozygous intercrosses were performed to confirm the embryonic lethality of the null allele. Mice heterozygous for this frataxin global knockout allele are available as Stock No. 028040.

The PV-Cre knock-in allele (also called Pvalb-IRES-Cre) was designed by Dr. Silvia Arber (Friedrich Miescher Institute) to have an IRES-Cre-pA targeting cassette (containing an internal ribosome entry site (IRES), a Cre recombinase coding sequence, a TK-Neo-Hygro sequence and polyadenylation site sequence) inserted into the 3' UTR of exon 5 of the parvalbumin (Pvalb) locus (see Stock No. 017320).

Allele

Symbol: Fxn^em2Lutzy
Name: endonuclease-mediated mutation 2, Cathy Lutz
MGI accession ID: MGI:5825257
Generation method: Endonuclease-mediated
Attributes: Conditional ready (e.g. floxed); No functional change
Marker symbol: Fxn
Marker name: frataxin
Chromosome: 19
Strain of origin: C57BL/6J
Molecular note: CRISPR/Cas9 genome editing is used to introduce loxP sites flanking exon 2.

Allele

Symbol: Fxn^em2.1Lutzy
Name: endonuclease-mediated mutation 2.1, Cathy Lutz
MGI accession ID: MGI:5825278
Generation method: Endonuclease-mediated
Attributes: Null/Knockout
Marker symbol: Fxn
Marker name: frataxin
Chromosome: 19
Strain of origin: C57BL/6J
Site of expression: Cardiac and skeletal muscle.
Molecular note: CRISPR/Cas9 genome editing was used to introduce loxP sites flanking exon 2. Cre-mediated recombination removed exon 2.

Allele

Symbol: Pvalb^tm1(cre)Arbr
Name: targeted mutation 1, Silvia Arber
MGI accession ID: MGI:3590684
Generation method: Targeted
Attributes: Recombinase-expressing
Marker symbol: Pvalb
Marker name: parvalbumin
Chromosome: 15
Strain of origin: 129P2/OlaHsd
Expressed genes: cre,cre recombinase,bacteriophage P1
Site of expression: most neurons that express parvalbumin including interneurons in the brain and proprioceptive afferent sensory neurons in the dorsal root ganglia
Molecular note: An IRES-cre-pA cassette was inserted into the 3' UTR of exon 5.