STOCK Mnx1^tm4(cre)Tmj Grm7^{Tg(SMN2)89Ahmb} Smn1^tm1Msd Tg(SMN2*delta7)4299Ahmb/J

Stock number: 007022
Research areas: Neurobiology Research

Description

Cre recombinase expression in these double targeted mutant mice is directed to motor neurons; mice exhibit symptoms and neuropathology similar to patients afflicted with severe proximal spinal muscular atrophy (SMA).

Detailed description

Mice homozygous for the Tg(SMN2*delta7)4299Ahmb and Tg(SMN2)89Ahmb transgenes and the Smn^tm1Msd targeted mutation allele exhibit symptoms and neuropathology similar to patients afflicted with severe proximal spinal muscular atrophy (SMA), and a similar phenotype observed in Stock no. 005025. At birth, triple mutants are noticeably smaller than normal littermates. By day 5, signs of muscle weakness are apparent and become progressively more pronounced over the following week as the mice display an abnormal gait, shakiness in the hind limbs and a tendency to fall over. In addition, this strain carries the Mnx1, HB9^cre targeted mutation with cre expression replacing HB9 (Hlxb9 or Mnx1) expression. Under control of the endogenous upstream elements, cre expression is directed to motor neurons. This strain can be used in conjunction with STOCK Smn1^{tm3(SMN2/Smn1)Mrph} Tg(SMN2*delta7)4299Ahmb Tg(SMN2)89Ahmb/J (Stock no. 007951), carrying the targeted mutation Smn1^{tm3(SMN2/Smn1)Mrph} allele which is engineered to revert to a fully functional Smn1 allele upon Cre-mediated recombination. Together these strains can be used to examine the effects of motor neuron expression of Smn1 in the pathology of Spinal Muscular Atrophy.

Development

This double targeted mutation, double transgenic strain was generated by crossing FVB.Cg-Tg(SMN2*delta7)4299Ahmb Tg(SMN2)89Ahmb Smn1^tm1Msd/J mice (Stock no. 005025) with B6.129S1-Mnx1^tm4(cre)Tmj/J mice (Stock no. 006600).



For the FVB.Cg-Tg(SMN2*delta7)4299Ahmb Tg(SMN2)89Ahmb Smn1^tm1Msd/J strain (Stock no. 005025): a targeted mutant allele was created in the laboratory of Dr. Michael Sendtner at the University of Wurzburg, Germany. Exon 2 of the endogenous mouse Smn gene was disrupted by employing a targeting vector encoding a neomycin cassette and a lacZ gene fused to the first 40 nucleotides of the disrupted exon to permit expression of the lacZ gene in tissues where Smn is normally expressed. The construct was electroporated into 129P2/OlaHsd-derived E14Tg2a-IV embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts and chimeric animals obtained. Chimeric animals were crossed to C57BL/6 for an unspecified number of generations.
The transgenic alleles were created in the laboratory of Dr. Arthur Burghes at Ohio State University. A 35.5 kb BamHI genomic fragment encoding the human SMN2 promoter and gene (derived from genomic clone PAC215P15) was injected into fertilized FVB/N mouse oocytes and founder animal 89 was obtained. Similarly, a human SMN2 cDNA (SMNdelta7) lacking exon 7 under the control of the human SMN2 promoter was microinjected into fertilized FVB/N oocytes and founder animal 4299 was obtained. Founder animal 89 was mated to mice heterozygous for the targeted mutation of the endogenous mouse Smn gene. These double mutants were in turn mated with mice bearing the SMNdelta7 transgenic allele. The triple mutant was then backcrossed to FVB/N for at least 6 generations.





For the B6.129S1-Mnx1^tm4(cre)Tmj/J strain (Stock no. 006600): a targeting vector containing an internal ribosome entry site (IRES)-Cre cassette, SV40 polyA sequence, and loxP-flanked pgk-neo cassette was designed to replace a portion of the first exon of the targeted gene. The construct was electroporated into 129S1/Sv-derived W9.5 embryonic stem (ES) cells. Recombinant clones were injected into C57BL/6J blastocysts to generate chimeric founders that transmitted the mutant allele. These HB9^cre mice were then backcrossed for at least 5 generations prior to arrival at The Jackson Laboratory.

Allele

Symbol: Tg(SMN2*delta7)4299Ahmb
Name: transgene insertion 4299, Arthur H M Burghes
MGI accession ID: MGI:3056918
Generation method: Transgenic
Attributes: Inserted expressed sequence; Humanized sequence
Marker symbol: Tg(SMN2*delta7)4299Ahmb
Marker name: transgene insertion 4299, Arthur H M Burghes
Chromosome: UN
Strain of origin: FVB/N
Expressed genes: SMN2,survival of motor neuron 2, centromeric,human
Site of expression: Tg(SMN2*delta7)4299Ahmb
Molecular note: The transgene contains a human SMN2 promoter and a human SMN2 cDNA (SMNdelta7) that lacks exon 7. There are 14 copies of this allele integrated into the genome.

Allele

Symbol: Mnx1^tm4(cre)Tmj
Name: targeted mutation 4, Thomas M Jessell
MGI accession ID: MGI:2447793
Generation method: Targeted
Attributes: Recombinase-expressing
Marker symbol: Mnx1
Marker name: motor neuron and pancreas homeobox 1
Chromosome: 5
Strain of origin: 129S1/Sv-Oca2⁺ Tyr⁺ Kitl⁺
Expressed genes: cre,cre recombinase,bacteriophage P1
Site of expression: motor neurons
Molecular note: Exon 1 was disrupted by the insertion of a cassette containing an IRES-cre gene, a floxed neomycin gene, and a SV40 polyadenylation signal.

Allele

Symbol: Smn1^tm1Msd
Name: targeted mutation 1, Michael Sendtner
MGI accession ID: MGI:2183946
Generation method: Targeted
Attributes: Reporter; Null/Knockout
Marker symbol: Smn1
Marker name: survival motor neuron 1
Chromosome: 13
Strain of origin: 129P2/OlaHsd
Expressed genes: lacZ,beta-galactosidase,E. coli
Site of expression: The expression of the lacZ gene in tissues where Smn is normally expressed was noted.
Molecular note: A lacZ-neo cassette was inserted into exon 2 by homologous recombination resulting in an in-frame fusion of lacZ to exon 2. Homozygous mutant embryos were identified up to 80 hours post coitum. The expression of the lacZ gene in tissues where Smn is normally expressed was noted.

Allele

Symbol: Grm7^{Tg(SMN2)89Ahmb}
Name: transgene insertion 89, Arthur H M Burghes
MGI accession ID: MGI:2448989
Generation method: Transgenic
Attributes: Hypomorph; Inserted expressed sequence; Humanized sequence
Marker symbol: Grm7
Marker name: glutamate receptor, metabotropic 7
Chromosome: 6
Strain of origin: FVB/N
Expressed genes: SMN2,survival of motor neuron 2, centromeric,human
Site of expression: Human SMN2 protein is expressed in the tissues examined (brain, liver, spinal cord) [Stock No. 005024].
Molecular note: A 35.5 kb genomic fragment containing the human survival motor neuron 2 (SMN2) gene and promoter was used for the transgene. The transgene is ubiquitously expressed in all tissues examined by Northern blot analysis. Line 89 carries 1 copy of the transgene integrated into intron 4 of the gene. RT-PCR confirmed reduced expression of the gene the transgene is integrated into.