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The targeting vector was constructed by inserting two contiguous PCR-amplified fragments from 129X1/SvJ genomic DNA (RRID:IMSR_JAX:000691) of the Gne gene to the pBluescript SK plasmid. The first 3.45-kbp fragment contained the last 3 exons of Gne, and the second fragment included 3.1 kbp downstream the Gne gene. The genetic variation predicting M743T was introduced by site-directed mutagenesis. The neomycin phosphotransferase gene (neo) flanked by LoxP sites and the diphtheria toxin A chain gene, each driven by the phosphoglycerate kinase (PGK) promoter was introduced into the vector as positive and negative selection markers, respectively. The PvuI-linearized targeting vector was electroporated into R1 embryonic stem cells. Eight correctly targeted clones (G418 selection) were identified by PCR using primers spanning from the neo gene to endogenous genomic sequences adjacent to the homologous recombination region. Two of the clones were aggregated with morulae-stage embryos to produce chimeras. Male chimeras were mated with ICR females to produce GneM743T/+ offspring, which were intercrossed to produce GneM743T/M743T mice. Subsequent PCR reconfirmed the genotype.
Sela I, Yakovlev L, Becker Cohen M, Elbaz M, Yanay N, Ben Shlomo U, Yotvat H, Fellig Y, Argov Z, Mitrani-Rosenbaum S. Variable phenotypes of knockin micecarrying the M712T Gne mutation. Neuromolecular Med. 2013 Mar;15(1):180-91. doi: 10.1007/s12017-012-8209-7. Epub 2012 Dec 13. (Medline PMID: 23238814)
Benyamini H, Kling Y, Yakovlev L, Becker Cohen M, Nevo Y, Elgavish S, HaraziA, Argov Z, Sela I, Mitrani-Rosenbaum S. Upregulation of Hallmark Muscle GenesProtects GneM743T/M743T Mutated Knock-In Mice From Kidney and Muscle Phenotype. JNeuromuscul Dis. 2020;7(2):119-136. doi: 10.3233/JND-190461. (Medline PMID: 31985472)
Cryo-recovered strains distributed by the MMRRC at JAX are shipped to the customer from the Pathogen & Opportunistic-Free Animal Room G200 - see https://www.jax.org/jax-mice-and-services/customer-support/customer-service/animal-health/health-status-reports.
Limited quantities of breeder mice (recovered litter) are available from a cryoarchive; recovered litter usually available to ship in 3 to 4 months.
Cryopreserved material may be available upon request, please inquire to csmmrrc@jax.org for more information.
The donor or their institution limits the distribution to non-profit institutions only.
Distribution of this strain requires submission of the MMRRC Conditions of Use (COU). A link to the COU web form will be provided via email after an order has been placed; the form should be completed then or the email forwarded to your institutional official for completion.
Additional charges may apply for any special requests. Shipping costs are in addition to the basic distribution/resuscitation fees. Information on shipping costs and any additional charges will be provided by the supplying MMRRC facility.
1 The distribution fee covers the expense of rederiving mice from a live mouse; you will receive the resulting litter. The litter will contain at minimum one mutant carrier; the actual number of animals and the gender and genotype ratios will vary. (Typically, multiple breeder pairs can be established from the recovered litter.) Prior to shipment, the MMRRC will provide information about the animals recovered. If you anticipate or find that you need to request specific genotypes, genders or quantities of mice in excess of what is likely from a resuscitated litter, you may discuss available options and pricing with the supplying MMRRC facility.
2 An aliquot contains a sufficient number of embryos (in one or more vials or straws and based on the transfer success rate of the MMRRC facility) to transfer into one to three recipients. The MMRRC makes no guarantee concerning embryo transfer success experienced in the recipient investigator's laboratory. Neither gender nor genotype ratios are guaranteed.
3 An aliquot is one straw or vial with sufficient sperm to recover at least one litter of mice, as per provided protocols, when performed at the MMRRC facility. The MMRRC makes no guarantee concerning the success of these procedures when performed outside the MMRRC facilities.
4 The distribution fee covers the expense of resuscitating mice from the cryo-archive; you will receive the resulting litter. The litter will contain at minimum one mutant carrier; the actual number of animals and the gender and genotype ratios will vary. (Typically, multiple breeder pairs can be established from the recovered litter.) Prior to shipment, the MMRRC will provide information about the animals recovered. If you anticipate or find that you need to request specific genotypes, genders or quantities of mice in excess of what is likely from a resuscitated litter, you may discuss available options and pricing with the supplying MMRRC facility.
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