Your gene of interest is inserted in a pENTR plasmid by RE cloning
Your gene of interest in inserted in a pENTR plasmid by PCR and Gateway cloning (BP reaction)
Your microRNA is designed by us, ordered for gene synthesis with 3 hairpins, then cloned in a pENTR plasmid by RE cloning
Your pENTR plasmid is cloned in a lentivector plasmid (monocistronic, bicistronic, autoinducible) by Gateway cloning (LR reaction). The final plasmid is maxipreped (average yield is 500 µg)
Your lentivector is produced, concentrated and aliquoted. Average yield. Average concentration.
|Lentivector titration by FACS||150||
Your lentivector is titrated on HT-1080 cells and by FACS (must contain a living color and a promoter active in HT-1080 cells)
|Lentivector titration by qPCR||200||In all other situations, your lentivector is titrated on HT-1080 cells and by qPCR.|