These Shh::gfp mice harbor a targeted mutation of the Sonic hedgehog (Shh) locus that secretes, after normal Shh processing, a bioactive GFP-tagged Shh signaling ligand (N-Shh::GFPp). These Shh::gfp may be useful to directly visualize the function of Shh ligand in an in vivo context, such as during neural tube patterning.
Andrew P McMahon, University of Southern California
While mice heterozygous for the Shh::gfp allele are viable, fertile, and indistinguishable from wild-type littermates, homozygotes are stillborn and show developmental defects consistent with reduced Sonic Hedgehog (Shh) signaling. The Shh::gfp mutation has GFP inserted into the endogenous Shh processing site (and adds a new processing site after the GFP). Thus normal Shh processing leads to secretion of the GFP-tagged Shh signaling ligand (N-Shh::GFPp) instead of wild-type Shh; with N-Shh::GFPp retaining both GFP and lipid modifications post-processing. Biochemical and cellular analysis indicates that Shh::GFP undergoes correct processing to produce active, bi-lipidated signaling peptides. Shh::GFP processing is, however, less efficient and results in reduced levels of Shh::GFP compared with wild-type Shh protein. These Shh::gfp mice produce bioactive, fluorescently labeled Shh from the endogenous Shh locus and may be useful to directly visualize the function of Shh ligand in an in vivo context, such as during neural tube patterning. In addition, these mice may also be useful in conjunction with other Sonic Hedgehog (Shh) mutant strains including Shh knockout mice (Stock No. 003318), Shh-floxed mice (Stock No. 004293), Shh-GFP/cre mice (Stock No. 005622), and Shh-creERT2 mice (Stock No. 005623).
A targeting vector was designed to replace the endogenous intein cleavage-cholesterol attachment site (Shh processing site) in exon 3 of the targeted gene with a Green Fluorescent Protein (GFP) followed by a new intein cleavage-cholesterol attachment site (Shh processing site), as well as insert a frt-flanked PGK-Neo cassette in intron 2. The donating investigator reports that this construct was microinjected into 129X1/SvJ-derived AV3 embryonic stem (ES) cells, correctly targeted ES cells were injected into recipient blastocysts, and chimeric mice were bred to C57BL/6 inbred mice to generate Shh::gfp mice. These Shh::gfp mice were reported to have been bred with C57BL/6 mice for a small number generations prior to sending to The Jackson Laboratory Repository in 2008 (see SNP notes below). Upon arrival, these mice were bred to C57BL/6J inbred mice (Stock No. 000664) to establish the colony. Over the next two years, the colony was backcrossed to C57BL/6J and/or wildtype mice from the colony for several additional generations. By March 2011, the colony was found to be ~96% C57BL/6 genetic background by SNP analysis (see SNP notes below).
In 2009, a 27 SNP (single nucleotide polymorphism) panel analysis performed on the rederived living colony at The Jackson Laboratory Repository. This revealed 16 markers (across several different chromosomes) that were not C57BL/6 allele-type. These data suggest the strain was sent to The Jackson Laboratory Repository with multiple sources of genetic background contamination (more than 129 genetic background).
In 2011, a 32 SNP (single nucleotide polymorphism) panel analysis, with markers covering all 19 chromosomes and the X chromosome, was performed on the colony at The Jackson Laboratory Repository. This revealed one marker on chromosome 1 (~89.1 Mbp) that was not fixed for C57BL/6 allele-type (e.g.: still segregating for 129X1 allele-type).
|Expressed Gene||GFP, Green Fluorescent Protein,|
|Site of Expression||early neuronal precursors|
|Allele Name||targeted mutation 6, Andrew P McMahon|
|Allele Type||Targeted (Reporter, Null/Knockout)|
|Allele Synonym(s)||Shhtm6Amc; targeted mutation 6, Andrew P McMahon|
|Gene Symbol and Name||Shh, sonic hedgehog|
|Gene Synonym(s)||Hhg1; short digits; M100081; Hx; Hxl3; HHG1; HPE3; Hx; Hxl3; SMMCI; HLP3; hedgehog gene 1; hemimelic extra toes; hemimelic extratoes like 3; TPT; TPTPS; MCOPCB5; Dsh; Hhg1; ShhNC|
|Expressed Gene||GFP, Green Fluorescent Protein,|
|Site of Expression||early neuronal precursors|
|Strain of Origin||Not Specified|
|Molecular Note||A GFP cassette was inserted into exon 3 and an frt-flanked neo cassette was inserted between exon 2 and 3. The presence of the fusion protein and the absence of the endogenous product was confirmed by western blot analysis on E9.5 embryo extracts. This allele produces functional albeit inefficient protein product.|
|Mutations Made By|| |
Andrew McMahon, University of Southern California
When maintaining a live colony, heterozygous mice may be bred together or to wildtype littermates.
When using the B6.129X1(Cg)-Shhtm6Amc/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #008466 in your Materials and Methods section.
|Heterozygous or Wild-type for Shh<tm6Amc>|
We will fulfill your order by providing at least two carriers for each strain ordered. The total number, sex, and genotypes provided will vary, although typically 8 or more animals are provided. Please check genotypes which will be recovered. While the genotypes of all animals produced will be communicated to you prior to scheduling shipment, the genotypes of animals provided may not reflect the mating scheme and genotypes described in the strain description. Animals are typically ready to ship in 11-14 weeks. If a second recovery is required to produce the minimum number of animals, then delivery time would increase to approximately 25 weeks. If we fail to produce animals of the correct genotype, you will not be charged. We cannot guarantee the reproductive success of mice shipped to your facility. If the mice are lost after the first three days (post-arrival) or do not produce progeny at your facility, a new order and fee will be necessary.
Cryorecovery to establish a Dedicated Supply for greater quantities of mice. Mice recovered can be used to establish a dedicated colony to contractually supply you mice according to your requirements. Price by quotation.
|Frozen Mouse Embryo||$2,595.00 per straw or vial|
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