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November 22 notes, part 23

November 22 notes, part 23 - General strategy for gene...

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Unformatted text preview: General strategy for gene targeting in mice Step 1 Gene targeting in ES cells E Inserted DNA 3 Veda neo' HSV-tk — a H l DNA H logoua DNA onto ogotn orno $553313, 2. Construction or targeting vector cells are wnm‘“ , The vector contains pieces of DNA that are homolo- lrom mouse ple- gone to the target gene. as wel as inserted DNA which "mum,“ embryos \ dungse the target gene and um lor positive- nsptive seledion ”mm“ ES cells 3. as cell mnmdion The celular machinery Ior homologous recombination allows the targeting vector enables the target vector to I W ‘\ €§ find and recombine with the target gene. ' .. W ‘1'”, HSV~tk Rare cel carryi target ed 99'“? "9 Target gene : 1 = ' 2—: Targets-Io \. warm 4. Prollleratlon A of MW ES cell Selection for preeeoce ol neo’ mdebeenceot HSV-tk enriches targeted Purepopulation of ES cells ES cells. curry-19 “"9““ gene 5. IllocIonoIEs commonwealth ThetargetedEScele ”Mmtheymixondiorrnornouic Theinjectedhhetooyotursinplmted seiniected witlttneoelooitneinnsroslmue mnemogmmothermethey ‘ trunwnidttneernbryodevslope. developinto moonicembryoe. Mouic inner col mess ‘ :l “autumn Born mosaic mice III II Thentoeeiomioematewitlr normdmioe &' a a to produce both gene targeted and nonnal all-m meumme Mfifififi- -°%n'i§* rnioo' when the Iergeudgono is mounted c mwmmn’mum Imam:mneou ...
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