Cloning Strategies
siRNA cloning strategies
If you have your siRNA (shRNA) already in pSUPER (or any other plasmid under control of PolIII promoter) you may use EcoR1-Cla1 sites to replace H1 promoter in pLVTH or pLVTHM with H1-shRNA cassette from pSUPER (or other plasmid).
BamH1-Cla1 cloning removes tetO element in pLVTH or pLVTHM that results in pLVH version
pLVTHM is a new version of pLVTH that allows for direct cloning of annealed shRNA into the lentiviral vector. In that case you would need to design your shRNA as depicted below.
Keep in mind that 5' of your antisense oligo starts at 5'CGATTTCCAAA.... (doh!)