pEco-Lenti-U6-shRNA-(RFP-Puro)CloningKitisdesignedtocloningofdouble-strandDNAencodingashorthairpinRNA.Oncetranscribed,theshRNAwasprocessedintoshortRNAinvivoforRNAianalysis.TomakeshRNAexpressingvector,twosyntheticoligonucleotideswerefirstannealedtoformthedoublestandDNAduplexwhichwasthenclonedintotheReady-to-use,linearpEco-Lenti-U6-(RFP-Puro)vectorviaT4enzymeligation.

TheexpressionofshRNAwasdrivenbyaconstitutivehumanU6promoter,aRNApolymeraseIIIpromoter.ThisKitprovidesmaterialsforgenerating10ofyourownshRNAexpressionlentivectors withthefollowingadvancedfeatures. Pleaseseeproductmanualfordetails.

shRNAvectorcloningscheme: 

Kitcontents:

pEco-Lenti-U6-(RFP-Puro)linearvector:                 10ul(10rxn,1ul/rxn)
10xshRNAoligoannealingsolution:                        50ul
5Xligationbuffer:                                                 50ul
T4ligaseenzyme:                                                 10ul(10rxn,1ul/rxn)
Cloningcontrolinsert:annealedLuc-shRNAduplex:   5ul(10rxn,1ul/rxn)
Sh-Sequencingprimer:                                          10ul(10xn,1ul/rxn)

Materialrequiredbutnotincluded:
DH5achemicalcompetentcells:                        10vials(10xn,1vial/rxn) 

KeyFeatures:

1. Linearizedvectorandreadyforuse,noneedforthetediousbenchworksforpreparationofvectorbackbone;