In-FusionCloningproductsprovidetheflexibilitytoperformsite-directedmutagenesis(deletions,basesubstitutions,oradditions),inadditiontopoweringanyofyoursingle-andmultiple-insertcloningexperiments.In-FusionCloningmakesiteasytoperformmutagenesisbycombiningthepoweroftheIn-FusionHDenzymewithinversePCR,amethodforrapidinvitroamplificationoftheDNAsequencesthatflankaregionwithaknownsequence.
In-FusionCloningproductsprovidetheflexibilitytoperformsite-directedmutagenesis(deletions,basesubstitutions,oradditions),inadditiontopoweringanyofyoursingle-andmultiple-insertcloningexperiments.In-FusionCloningmakesiteasytoperformmutagenesisbycombiningthepoweroftheIn-FusionHDenzymewithinversePCR,amethodforrapidinvitroamplificationoftheDNAsequencesthatflankaregionwithaknownsequence.
DuringinversePCR,primersareorientedinoppositedirectionsonyourcircularcloningvector,sotoperformmutagenesiswithIn-Fusionsystems,designyourPCRprimerssothattheyhavea15-bpoverlapwitheachotherattheir5"endsandincorporatethemutationofinterest.UsetheCloneAmpHiFiPCRPremix(ahigh-fidelityPCRpolymeraseincludedwithallIn-FusionHDCloningPlussystems)toperformyourPCRreaction,addtheIn-FusionHDenzymepremixtoyourlinearizedPCRproduct,andtransformintotheprovidedStellarCompetentCells.Youhavea≥95%chanceofrecoveringyourfinaldesiredconstruct—thefirsttimeandeverytime.
