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Alabama Neuroscience Blueprint Core B- Molecular Engineering

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Alabama Neuroscience Blueprint Core B- Molecular Engineering

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    2. Use of genetically modified mice in biomedical research

    3. Core B Function One of the greatest obstacles for investigators who wish to use mouse genetics to study their gene of interest is in the generation of DNA-targeting constructs. The Molecular Engineering Core will function to generate targeting cassettes for the production of both transgenic and knockout animals *(Core B does not perform ES electroporation, blastocyst injections or DNA microinjections- see Dr. Bob Kesterson at the UAB transgenic mouse facility for these services) To start a project, go to website and fill out job request form

    4. Core B Services Advise participating investigators on the generation and design of DNA constructs that are optimal for their studies Construct the necessary targeting cassettes and prep DNA for injections or electroporation 3. Assist in the development of genotyping protocols for the maintenance of new mouse lines.

    5. Transgenic Constructs

    6. Constitutive gene-inactivation constructs Gene-trap alleles Constitutive inactivation Tagged with B-gal for use in determining expression patterns ES cells libraries already generated database search identifies possible targeted genes Bay genomics/MMRRC allele may be null or hypomorphic

    7. Constitutive gene-inactivation constructs Constitutive gene knockout Production of gene-replacement cassettes Identify BACs encoding gene Retrieve genomic fragments and replace “exons” with the ES cell selectable marker neomycin Sequence construct to confirm correct replacement Prep DNA for electroporation into ES cells

    8. Conditional gene-inactivation constructs Conditional gene knockout using Cre-lox methodology Two components insertion of loxP sites into a “neutral” region surrounding critical exon/s expression of Cre recombinase Allows for gene inactivation in a restricted neuronal population and at desired time in development Construct floxed allele Sequence confirm and prep DNA for electroporation

    12. Genotyping The core will develop PCR based genotyping protocols for DNA targeting cassettes Both standard PCR and qPCR genotyping available Provide assistance and information on maintenance of mouse colony breeding, colony size and maintenance, book keeping tail clip, ear punch, mating, weaning, chow

    13. Core B- Molecular Engineering Personnel Open Position- Technical Director Sylvia Huang- Research Assistant Ludwig Francillion- Laboratory Assistant Contact Info Scott Wilson 5-5573 SHELBY 972

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