B6.129P2-Pitx3^tm1Mli/Mmjax

Stock number: 028554
Product name: Pitx3-GFP
Strain synonyms: Pitx3-GFP
Research areas: Developmental Biology Research, Neurobiology Research, Research Tools, Sensorineural Research

Description

Homozygous Pitx3^GFP knockin/knockout mice exhibit a progressive loss of dopaminergic neurons in the midbrain as well as an absence of lens epithelial cells. These mice may be useful in studying in vivo imaging, fate mapping of dopaminergic neurons in the midbrain and lens development.

Detailed description

The Pitx3 gene encodes a homeobox transcription factor that is involved in dopaminergic neuron development and lens formation during eye development. Mutations in Pitx3 are associated with cataract. The Pitx3-GFP knockin/knockout allele has an SA-eGFP-IRES-pac-pA cassette replacing part of intron 1, exons 2-3 and half of exon 4. Pitx3-directed GFP fluorescence is observed in the dopaminergic neurons in the substantia nigra and ventral tegmental area of the adult midbrain. Mice homozygous for the mutation exhibit a progressive loss of dopaminergic neurons in the midbrain as well as a deformed lens due to a depletion of lens epithelial cells and aberrant fiber cell differentiation. These mice may be useful in studying in vivo imaging, fate mapping of dopaminergic neurons in the midbrain and lens development.

Development

A targeting vector was designed by Dr. Meng Li (University of Edinburgh) to replace part of intron 1, exons 2-3 and half of exon 4 of the paired-like homeodomain transcription factor 3 gene (Pitx3) on chromosome 19 with an SA (splice acceptor)-eGFP-IRES-pac (puromycin)-pA cassette followed by a loxP-flanked PGK-hygromycin cassette. The construct was electroporated into 129P2/OlaHsd-derived E14TG2a embryonic stem (ES) cells. One of the recombinant clones was electroporated with a Cre-expressing vector to remove the hygromycin cassette. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6 females to establish the colony. The resulting Pitx3^GFP mice were bred with C57BL/6 mice for at least 15 generations prior to sending males to The Jackson Laboratory Repository in 2015. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).

A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 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. While the 27 markers throughout the genome suggested a C57BL/6 genetic background, all 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 C57BL/6N genetic background.

Allele

Symbol: Pitx3^tm1Mli
Name: targeted mutation 1, Meng Li
MGI accession ID: MGI:3573869
Generation method: Targeted
Attributes: Reporter; Null/Knockout
Marker symbol: Pitx3
Marker name: paired-like homeodomain transcription factor 3
Chromosome: 19
Strain of origin: 129P2/OlaHsd
Expressed genes: GFP,Green Fluorescent Protein,
Site of expression: GFP fluorescence is observed in the dopaminergic neurons in the substantia nigra and ventral tegmental area of the adult midbrain.
Molecular note: A targeting vector containing a splice acceptor-eGFP-ires-pac-pA cassette and a floxed PGK-hygro cassette was used to replace exons 2, 3 and half of 4. Expression of a Cre plasmid excised the floxed hygro.