STOCK Tg(Chx10-EGFP/cre,-ALPP)2Clc/J

Stock number: 005105
Product name: Chx10 BAC
Research areas: Neurobiology Research, Research Tools

Description

These Chx10 BAC mice express a transgene containing EGFP and human placental alkaline phosphatase with reporter activity being seen in a subset of Müller glial cells.

Detailed description

Mice hemizygous for the transgenic insert are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. The transgene insert contains a fusion product involving Cre recombinase and an Enhanced Green Fluorescent Protein (EGFP) and a fusion product involving an internal ribosome entry site/human placental alkaline phosphatase under the control of the mouse Chx10, C. elegans ceh-10 homeo domain containing homolog promoter. This strain serves as a multifunctional reporter strain. Expression of the Chx10 BAC, as detected by in situ hybridization, mimics the endogenous Chx10 gene expression pattern. Alkaline phosphatase expression is mosaic, but specific to the retina and Muller glial cells. EGFP expression is detected in the outer neuroblastic layer of the retina at embryonic days 14.5, 17.5 and neonates. When crossed to a cre reporter strain, the resulting mice exhibit mosaicism in reporter gene expression. This mutant mouse strain may be useful in studies of retinal progenitor and bipolar cell lineage or fate mapping.

Development

A transgenic construct encoding an EGFP/cre fusion protein, an internal ribosome entry site/human placental alkaline phosphatase fusion protein and an FRT-flanked selection cassette (composed of neomycin resistance and kanamycin resistance genes under the control of phosphoglycerate kinase and Tn5 transposon promoter), was inserted into a BAC encoding the mouse Chx10, C. elegans ceh-10 homeo domain containing homolog promoter. This modified BAC transgene was microinjected into B6;SJL fertilized eggs. The founder animals were bred to FVB. The mice were crossed to 129S4/SvJaeSor-Gt(ROSA)26Sor^tm1(FLP1)Dym/J mice (see Stock No. 003946) to remove the FRT-flanked selection cassette, and then later crossed to C57BL/6 for one generation.

Allele

Symbol: Tg(Chx10-EGFP/cre,-ALPP)2Clc
Name: transgene insertion 2, Constance L Cepko
MGI accession ID: MGI:3052237
Generation method: Transgenic
Attributes: Recombinase-expressing; Reporter
Marker symbol: Tg(Chx10-EGFP/cre,-ALPP)2Clc
Marker name: transgene insertion 2, Constance L Cepko
Chromosome: UN
Strain of origin: C57BL/6 and SJL
Expressed genes: cre,cre recombinase,bacteriophage P1; GFP,Green Fluorescent Protein,; ALPP,alkaline phosphatase, placental,human
Site of expression: expression pattern is mosaic, but specific to the retina and Muller glial cells
Molecular note: This reporter gene expresses an enhanced green fluorescent protein-Cre recombinase fusion protein and alkaline phosphatase under control of Chx10 promoter and enhancer elements.The transgene was generated in a 129/Sv-derived bacterial artificial chromosome (BAC) containing the Chx10 gene and extending approximately 55 kb upstream of the translation start site and about 22 kb downstream of the polyadenylation signal. The 8th codon of Chx10 is joined in-frame to an EGFP-cre fusion gene followed by an internal ribosome entry sequence (IRES) coupled to the human placental alkaline phosphatase gene; 32 codons are deleted from within the N-terminal coding sequence of Chx10. The transgene was found by Southern blot analysis to be present in multiple copies, most or all of them intact. The mice were crossed to 129S4/SvJaeSor-Gt(ROSA)26Sortm1(FLP1)Dym/J mice to remove the FRT-flanked selection cassette.
General note: According to Rowan and Cepko (2004), the modified BAC DNA "was injected into male pronuclei of SJL/B6 fertilized eggs."

Two transgenic lines were generated. Line 1 exhibited highly mosaic expression of the EGFP reporter. Line 2 expressed EGFP more uniformly, but weakly; excision of the FRT-neo cassette increased EGFP expression in subsequent generations, so line 2 was used for further studies.

Hemizygous transgenic mice are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. Alkaline phosphatase expression is mosaic, but specific to the retina, and is also detected in Muller glial cells. Green Fluorescent Protein (GFP) expression is detected in the outer neuroblastic layer of the retina at embryonic days 14.5, 17.5 and neonates. When crossed to a cre reporter strain, the resulting mice exhibit mosaicism in reporter gene expression.