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《pglo transformation lab ap lab 7 - brookings …：pglo轉(zhuǎn)化實(shí)驗(yàn)室ap實(shí)驗(yàn)室7 -布魯金斯…》由會(huì)員上傳分享，免費(fèi)在線閱讀，更多相關(guān)內(nèi)容在教育資源-天天文庫(kù)。
1、pGLOTransformationLAB APLAB6http://www.mshri.on.ca/nagy/GFP%20mice.jpgBIO-RADlabbookpGLOoriblaGFParaCAequoreavictoria:Sourceof“glowinggene”forthisexperimentJellyfishGeneputintoOtherCrittershttp://www.technologyreview.com/files/21291/monkey_x600.jpghttp://www.lafuga.de/GFP_pig.jpgPLASMIDExtrachrom
2、osomalDNAOftencarrygenesforantibioticresistanceCanbepassedfromonebacteriumtoanotherhttp://www.agen.ufl.edu/~owens/age2062/OnLineBiology/OLBB/www.emc.maricopa.edu/faculty/farabee/BIOBK/14_1.jpgBacterialTransformationPlasmidsChromosomalDNABacterialCellTheuptakeofDNApGLOLABSUPPLIESFOAMtubeholder/flo
3、at4-fliptopmicrotubesBlue-Transformingsolution(CaCl2)Yellow-LBnutrientbrothPink-label+Purple-label-1-colorederaser(toIDyourtubesinwaterbath)1-pkgyellowinnoculatingloops2-Sterilepipettes4pouredagarplates1-LB2-LB/amp1-LB/amp/araPERMANENTMARKERCupwithcrushediceLABELTUBESPurple=+pGLOpink=-pGLOUsester
4、ilepipettetoadd250μLtransformationsolutiontopGLO+and–tubesTransformation solution(CaCl2)GetyourrackonICE!INNOCULATETUBESWITHE.coliBACTERIAPickonecolony Twirlloopin+pGLOtubeGetnewloop Pickonecolony Twirlloopin–pGLOtubeUSESPECIAL GARBAGEBAGFOR DISPOSALOFUSEDLOOPSEXAMINEpGLOplasmidDNAUseUVlighttoexa
5、minepGLOplasmidvialUVlightcanbeharmfultoyoureyes! Wearyourgoggles. Donotshineineyes.GFP=GreenFluorescentProteinisolatedfromjellyfishUSEDASAGENETICTOOLhttp://www.mshri.on.ca/nagy/GFP%20mice.jpgPLASMIDDNATRANSFERTHISSTEPISCRUCIAL!Lookcloselytomakesureyouhaveafilmofsolutionacrossthering.(Similartoso
6、apyfilmwhenyoublowbubbles)ADDPLASMIDTO+TUBEDONOTADDPLASMID TO-TUBEPutrackonICEfor10MIN!WHILEYOURTUBESCOOLLABELYOURPLATESFLIPUPSIDEDOWNANDWRITELABELSONBOTTOM…NOTONTOP!LB(LuriaandBertani)–broth&agarprovidesnutrientsforbacterialgrowthLB/ampLuriaagar+ampicillin(antibiotic)LB/amp/araLuriaagar+ampicill
7、in+arabinosesugarSHOCKINGINCREASESUPTAKEOFFOREIGNDNA(PLASMID)OSMOTICSHOCK=TransformingsolutionCaCl2HEATSHOCKRAPIDTEMPERATURECHANGEisthekey50SECONDS2MINUTESPlacefoamrackwith+and–tubesondesktopUsenewsterilepipettetoadd25
