B6.Cg-Tg(Pitx3-IFP,-tdTomato,-Citrine)2Past/Mmjax

Stock number: 038153
Product name: Pitx3-ITC
Research areas: Neurobiology Research, Research Tools

Description

Pitx3-ITC transgenic mice constitutively express an IFP (infrared fluorescent protein) in substantia nigra pars compacta (SNc) midbrain dopaminergic neurons (mDA). In addition, this strain expresses a cre-dependent tdTomato reporter and Flp-dependent Citrine reporter. In this model, conditional reporter expression can be used as an intersectional genetic tool to analyze substantia nigra development and neuron migration in vivo.

Detailed description

Pitx3 (paired-like homeodomain transcription factor 3) encodes a transcription factor involved in dopaminergic neuron differentiation. The Pitx3 promoter restricts expression to substantia nigra pars compacta (SNc) midbrain dopaminergic neurons (mDA) from E11.5 onward.  Pitx3-ITC transgenic mice constitutively express an IFP (infrared fluorescent protein) under the direction of the Pitx3 promoter. cre-mediated recombination removes the IFP reporter and allows expression of the tdTomato reporter. Flp-mediated recombination removes IFP and tdTomato and allows expression of the Citrine reporter. Each reporter is followed by a stop signal that prevents expression of the next reporter. Immunohistochemistry combined with a marker (tyrosine hydroxylase, TH) for mDA neurons confirms expression in the lateral, but not medial, mDA neurons in adult mice crossed to cre or Flp-expressing strains. Mice hemizygous and males homozygous for the transgene are viable and fertile, however, homozygous mice are blind. Female homozygotes have not been tested for fertility. In this model, conditional reporter expression can be used as an intersectional genetic tool to analyze substantia nigra development and neuron migration in vivo.

Development

The Pitx3-ITC transgenic construct contains a BAC (RP23-125F3) modified to insert the ITC cassette into ATG start codon in exon 2 of the Pitx3 (paired-like homeodomain transcription factor 3) locus. The ITC construct includes: an FRT site, a loxP-flanked IFP (infrared fluorescent protein)/Sv40 polyA, a tdTomato/bGH polyA sequence, a second FRT site and a Citrine/synthetic polyA sequence. The linearized construct was microinjected into the pronuclei of fertilized oocytes created by mating (C57BL/6 x SJL)F1 or C57BL/6 females to (C57BL/6 x SJL)F1 males. Founder line 2 was established. The donating investigator backcrossed the strain to C57BL/6 for 10 generations. 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).

Allele

Symbol: Tg(Pitx3-IFP,-tdTomato,-Citrine)2Past
Name: transgene insertion 2, Jeroen Pasterkamp
MGI accession ID: MGI:6726603
Generation method: Transgenic
Attributes: Conditional ready (e.g. floxed); Reporter
Synonyms: Pitx3-ITC
Marker symbol: Tg(Pitx3-IFP,-tdTomato,-Citrine)2Past
Marker name: transgene insertion 2, Jeroen Pasterkamp
Chromosome: UN
Strain of origin: C57BL/6 x SJL
Molecular note: The transgene consists of mouse Pitx3 BAC clone RP23-125F3 with a ITC cassette inserted into the ATG start codon in exon 2 of the Pitx3 gene. The ITC cassette has the following components in order, FRT, loxP, IFP(infrared fluorescent protein)-polyA, loxP, tdTomato-polyA, FRT, Citrine-polyA, with each gene followed by a stop signal that prevents expression of the next gene. Without recombination, IFP is expressed. While Cre/loxP recombination will express tdTomato and Flp/frt recombination will express Citrine. Founder information and line numbers were not available. Different founders displayed comparable labeling patterns. Line 2 was used for the study.