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oreYouBegin PrecautionsNLysis/BindingBufferandWashBufferIcontainguanidinehydrochloride whichisanirritant.Alwayswearglovesandfollowstandardsafetypre- cautionstominimizecontactwhenhandling. ?Donotletthesebufferstouchyourskin,eyes,ormucousmembranes.If contactdoesoccur,washtheaffectedareaimmediatelywithlargeamounts ofwater;otherwise,thereagentmaycauseburns.Ifyouspillthereagent, dilutethespillwithwaterbeforewipingitup. ?NeverstoreorusetheBindingBuffernearhumanoranimalfood. ?Alwayswearglovesandfollowstandardsafetyprecautionswhenhandling thesebuffers. SampleMaterialSolidtissue(e.g.,mouseliver,spleen,lung,heart),1–10mg(formortar/pestle disruption)or1–25mg(forrotor-statorhomogenization). Handlingand Storageof StartingMaterial ToavoidanydegradationofRNAinsamplematerialbyintracellularRNases followtheseguidelines: ?Processthesamplematerialassoonascollected ?Ifstorageofsamplematerialisrequired,flashfreezeinliquidnitrogenand storeat-60°C NDonotallowfrozentissuematerialtothawduringhandling(e.g., weighing).Therelevantproceduresshouldbecarriedoutasquicklyas possible. ?Samplescanalsobestoredat-60°CorbelowinLysis/BindingBufferafter disruptionandhomogenization. LYieldsmayvarydependingonstoragetime. NUsesteriledisposablepolypropylenetubesandtipsinordertoavoid RNasecontamination.Wearglovesduringtheassay. Disruptionand Homogenization ?Efficientdisruptionandhomogenizationofthestartingmaterialisessential forintra-cellularRNAisolationproceduresfromtissues. ?Thecompletedisruptionofcellwallsandplasmamembranesofcellsand organellesisabsolutelyrequiredtoreleasealltheRNAcontainedinthesam- ple.Incompletedisruptionresultsinsignificantlyreducedyields. ?Homogenizationisnecessarytoreducetheviscosityofthecelllysatespro- ducedbydisruption.Homogenizationshearsthehighmolecularweight genomicDNAandotherhighmolecularweightcellularcomponentstocre- ateahomogeneouslysate. ?Incompletehomogenizationresultsinsignificantlyreducedyields.Somedis- ruptionmethodssimultaneouslyhomogenizethesample,whileothers requireanadditionalhomogenizationst 发自小木虫Android客户端 |
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