These SncaΔflox Rosa26-stop-lacZ reporter mice carry a knock out allele of the Snca gene and the Gt(ROSA)26Sortm1Sho Cre reporter allele. This strain may be useful for generating mutants in which Cre recombinase efficiency can be monitored for applications related to neurodegenerative synucleinopathies, such as Parkinson's disease and multiple system atrophy.
Vladimir L Buchman, Cardiff University School of Biosciences
Genetic Background | Generation |
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Allele Type | Gene Symbol | Gene Name |
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Targeted (Conditional ready (e.g. floxed), Reporter) | Gt(ROSA)26Sor | gene trap ROSA 26, Philippe Soriano |
Allele Type | Gene Symbol | Gene Name |
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Targeted (Null/Knockout) | Snca | synuclein, alpha |
These mice carry the SncaΔflox allele (028559) and the Rosa26-stop-lacZ reporter allele (003504).
The "SncaΔflox" knock-out allele contains a 1164 bp fragment deletion (including the first coding exon (exon II) and adjacent flanking sequence) of the Snca (synuclein, alpha) gene.
The Gt(ROSA)26Sortm1Sho allele contains a loxP-flanked DNA STOP sequence preventing expression of the downstream lacZ gene. When crossed with a cre recombinase expressing strain, the STOP sequence is removed and lacZ is expressed in cells/tissues where cre is expressed.
The Rosa26-stop-lacZ reporter allele in this double mutant line allows for the monitoring of Cre recombination efficiency when crossed to mice carrying a floxed Snca allele (see 025636).
Of note, because the Snca and Rosa26 loci are located on chromosome 6, approximately 50 Mb apart, there is a possibility that recombination can separate the mutant alleles during breeding.
Mice that are homozygous for these alleles are viable and fertile.
Mutations in human SNCA that result in aberrant polymerization and fibrillar aggregations are associated with neurodegenerative synucleinopathies, such as Parkinson's disease and multiple system atrophy.
Mice carrying SncaΔflox and ROSA26-stop-lacZ reporter (Proc Natl Acad Sci U S A. 1999 Apr 27;96(9):5037-42) loci linked at the same chromosome as the result of natural crossover were selected from a pool of animals generated during crossbreeding of two mouse lines (both on C57BL/6 background), each bearing one of these modified loci.
For the Sncatm1.2Vlb allele, a targeting vector containing a FRT site-flanked NEO cassette and a loxP site was inserted downstream of exon II (the first coding exon) of the targeted Snca gene, and another loxP site was inserted upstream of exon II. This construct was electroporated into C57BL/6N-Atm1Brd-derived JM8A3.N1 embryonic stem (ES) cells. The resulting chimeric animals were bred to C57BL/6J mice for 2 generations and then crossed to transgenic mice (on a congenic C57BL/6 genetic background) expressing FLP recombinase under the control of the human ACTB promoter, generating SncafloxΔneo mice. The FLP recombinase mediated excision of the FRT flanked NEO cassette and a 400 bp fragment encompassing a simple repeat region does not affect Snca locus expression. The SncafloxΔneo mice that retained the floxed exon II were bred to C57BL/6J mice for 2 generations to remove the FLP recombinase transgene, before being intercrossed to generate homozygotes. These homozygous SncafloxΔneo/floxΔneo mice were then bred to transgenic mice (on the C57BL/6 background) expressing Cre recombinase under the control of the CMV promoter (see, for example, Stock No. 006054) to excise the floxed exon II and flanking sequences. The resulting knock-out SncaΔflox mice were then bred to C57BL/6J to remove the Cre recombinase transgene and further backcrossed with C57BL/6J mice for more than 4 generations.
For the Gt(ROSA)26Sortm1Sho, the R26R targeting construct was created by subcloning the pROSA26-1 vector, and inserting a splice acceptor sequence (identical to the one used in the original gene trap allele), a loxP-flanked STOP sequence (containing a neo cassette and triple polyadenylation (STOP) sequence), a lacZ gene, and a polyadenylation sequence at a unique site approximately 300-bp 5' of the original gene trap integration site. Transcriptional read-through is prevented with the use of a triple polyadenylation (STOP) sequence at the 3' end of the neo expression cassette. The construct was electroporated into 129S4/SvJaeSor derived AK7 embryonic stem (ES) cells. (see, for example, Stock No. 003474)
The single mutant lines were crossed to generate double mutant animals. The mice were then bred to select for these SncaΔflox ROSA26-stop-lacZ reporter mice carrying both mutant alleles on the same chromosome.
The mice were then bred to homozygosity for both alleles.
Upon arrival, embryos were cryopreserved.
A 48 SNP (single nucleotide polymorphism) panel analysis, with 43 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. Two of the 27 markers throughout the genome were segregating, suggesting an incomplete backcross. One of 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 mixed C57BL/6J ; C57BL/6N genetic background.
Expressed Gene | Bgeo, fusion of beta-galactosidase and neomycin phosphotransferase genes, E. coli |
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Site of Expression | when crossed to a cre recombinase-expressing strain, lacZ expression is observed in the cre-expressing tissues; this is similar to Gt(ROSA)26Sortm1Sor but has one copy of the loxP-flanked reporter gene |
Site of Expression |
Allele Name | targeted mutation 1, Stuart Orkin |
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Allele Type | Targeted (Conditional ready (e.g. floxed), Reporter) |
Allele Synonym(s) | gtROSA; R26fslz; R26-beta-galfl-STOP; R26R; R26Rosa-lox-Stop-lox-LacZ; R26RstoplacZ; Rosa26fsLz; ROSA26-loxP-stoploxP-beta-geo; ROSA26R-LacZ reporter; ROSA26-stop-lacZ |
Gene Symbol and Name | Gt(ROSA)26Sor, gene trap ROSA 26, Philippe Soriano |
Gene Synonym(s) | |
Expressed Gene | Bgeo, fusion of beta-galactosidase and neomycin phosphotransferase genes, E. coli |
Site of Expression | when crossed to a cre recombinase-expressing strain, lacZ expression is observed in the cre-expressing tissues; this is similar to Gt(ROSA)26Sortm1Sor but has one copy of the loxP-flanked reporter gene |
Strain of Origin | 129S4/SvJaeSor |
Chromosome | 6 |
General Note | Mice carrying this mutation are considered to be reporter strains, with successful Cre excision indicated by beta-gal expression in Cre-expressing tissues. |
Molecular Note | A targeting vector was designed to insert floxed (flanked by loxP) phosphoglycerate kinase-1-cytosine deaminase/phosphoglycerate kinase-1-puromycin (PGK-CD/PGK-Puro) and stop sequences between the splicing acceptor (SA) and the beta-gal-neomycin phosphotransferase fusion gene (beta-geo) sequences of the proviral ROSA 26 locus. Correctly targeted, this resulted in ES cells that were Puro resistant but G418 susceptible, and did not express beta-gal. |
Mutations Made By | Dr. Xiaohong Mao, Novartis |
Allele Name | targeted mutation 1.2, Vladimir L Buchman |
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Allele Type | Targeted (Null/Knockout) |
Allele Synonym(s) | Sncadelta floxed |
Gene Symbol and Name | Snca, synuclein, alpha |
Gene Synonym(s) | |
Strain of Origin | C57BL/6N-Atm1Brd |
Chromosome | 6 |
Molecular Note | A targeting vector containing a FRT site-flanked neo cassette and a loxP site was inserted downstream of exon 2 (the first coding exon) of the targeted gene, and another loxP site was inserted upstream of exon 2. Flp-mediated recombination removed the FRT-flanked neo cassette leaving exon 2 floxed. Germline, Cre-mediated recombination removed exon 2. Western blot analysis of neural tissues confirmed that no protein is expressed. |
When maintaining a live colony, these mice can be bred as homozygous for both alleles. Of note, because the Snca and Rosa26 loci are located on chromosome 6, approximately 50 Mb apart, there is a possibility that recombination can separate the mutant alleles during breeding.
When using the Snca delta flox ("clean KO") & Rosa26-stop-lacZ mouse strain in a publication, please cite the originating article(s) and include JAX stock #028560 in your Materials and Methods section.
Facility Barrier Level Descriptions
Service/Product | Description | Price |
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Homozygous for Snca<tm1.2Vlb> and Gt(ROSA)26Sor<tm1Sho>, 1 pair minimum |
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The Jackson Laboratory has rigorous genetic quality control and mutant gene genotyping programs to ensure the genetic background of JAX® Mice strains as well as the genotypes of strains with identified molecular mutations. JAX® Mice strains are only made available to researchers after meeting our standards. However, the phenotype of each strain may not be fully characterized and/or captured in the strain data sheets. Therefore, we cannot guarantee a strain's phenotype will meet all expectations. To ensure that JAX® Mice will meet the needs of individual research projects or when requesting a strain that is new to your research, we suggest ordering and performing tests on a small number of mice to determine suitability for your particular project. We do not guarantee breeding performance and therefore suggest that investigators order more than one breeding pair to avoid delays in their research.
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