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TARGETING CONSTRUCT

f1 origin. A. p NosTerFRT-neo (5500bp). Amp r. Ori. Bgl II Xba I Not I Sac I Sfi I Fse I Avr II …… Not I Xba I Not I Dsa I Sac I. FRT. FRT. Nos-ter. Tn5. pGK. neo. 0.3kb. 1.8kb. UPSTREAM MULTIPLE CLONING SITE

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TARGETING CONSTRUCT

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  1. f1 origin A pNosTerFRT-neo (5500bp) Ampr Ori BglII XbaI NotI SacI SfiI FseIAvrII……NotI XbaI NotI DsaI SacI FRT FRT Nos-ter Tn5 pGK neo 0.3kb 1.8kb UPSTREAM MULTIPLE CLONING SITE NotI XbaI KnpI SacI MluINheIXhoIBglII HindIII….. BglII EcoRI PstI SmaIBamHISalIPstI B TARGETING CONSTRUCT SpeI/XhoI fragment from pST33 was cloned into pNosTerFRT-Neo cut with NheI/SalI GA Pc2ox1 (i) pGAPc2ox1FRT-neo EcoRI EcoRI EcoRVNotI 0.9kb BamHI/SacI fragment from pBin-mgfp5-ER was blunt end ligated into pNosTerFRT-Neo cut with SmaI mgfp5-ER (ii) pmgfp5-ERFRT-neo 0.8kb

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