A9E583D2d01 - Part II Mutagenesis Introduction Transgenic...

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Part II Mutagenesis
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Introduction Transgenic technology > genetic variation: - essential in research function gene “study abnormal to understand normal” but also - models for human diseases - process of positional cloning (searching responsible gene for certain phenotype) - chromosome engineering (trisomies, translocations, deletions,…) - life stock: disease resistant animals, faster growing animals, better meat ...
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Introduction Genetic variation: Introducing genes (constitutively, cell specific,…) Switch-off genes (constitutively, point mutants, …) More subtle: introducing point mutations, promotor research,. .. In general, to achieve by Transiënt systems Permanent systems
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Transiënt systems (= gene therapy) Adenovirusses Retrovirusses/Lentivirusses Adeno Associated Virus nude cDNA +/- integration 1 generation only in somatic cells > not in the germline > not inheritable Not in this course
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Introduction genetic variation: spontanous mutagenesis very seldom in mouse (1/10 5 spermatozoa has visual mutation) via genetic manipulation > generate mutants “trangenic animal s.l.” :all mutants that can be achieved by intervention of man and that could ‘impossibly’ arise spontanously in nature.
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Introduction genetic variation: Some examples of spontaneous mutations in mice: Brachyury Extra toes (Xt) nude Small eye
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1980 1990 2000 Pronucleus injection in zygotes Alternatives for pronucleus injection ES cell technology For expression of Foreign gene via Transgenic way ES cell technology Alternatives for ES For elimination of own gene via Transgenic way
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Year Pubmed „Transgenic mouse“ 1982 11 1985 92 1988 542 1991 2206 1994 5537 1997 11853 2000 21079 2002 28692
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Very General: 1. Bringing in foreign genes in early embryo’s > mutant animals 70ies and 80ies till now especially via zygote’s 2. Modification of the genome of ES cells > chimeric embryo’s > mutant animals since end 80ies gene targeting gene trapping site specific recombination 3. Random mutagenesis/ENU
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5. Transgenic animals (overexpression “foreign” gene) - introduction DNA in mammals via embryo - why embryo ? Big chance for inheritance of the transgene
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number of essential technical demands: - take out pre-implantation embryo - culture in vitro of embryo (cf. oviduct-uterus) - introduction foreign genetic material in reproducible way (very invasive, life threatening, small cells) - re-introduction embryo + development to adult
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1970ies : first attempts to bring in foreign DNA in embryo’s 1. Teratocarcinoma cells transfect + introduce cells in host embryo > chimeric embryo > chimeric mouse but no “germline transmission”
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This note was uploaded on 05/30/2010 for the course WE CHBIBI0000 taught by Professor Krisvleminckx during the Spring '10 term at Ghent University.

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A9E583D2d01 - Part II Mutagenesis Introduction Transgenic...

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