Lenti tet on advanced inducible expression system

For the transfection of the HPCOA07/03, the modified Lenti-TM Tet-On^® Advanced Inducible Expression System (Clonetech) was used. To generate and package this virus, the envelope plasmid pMDG:VSV-G and the packaging plasmid P8.91:GAG-POL were used instead of the Lenti-X HT packaging mix, and the transfection was conducted using Lipofectamine instead of Lentiphos HT, as this setup was already in place in the lab. To generate the two viruses, one with the regulator vector and one with the response vector, HEK cells were transfected using Lipofectamine following the manufacturer’s instructions, with a total of 30 μg of DNA containing the envelope plasmid pMDG:VSV-G, the packaging plasmid P8.91:GAG-POL, and either the regulator or the response plasmid. The virus was collected 24 and 48 h after the transfection and concentrated using the Lenti-X concentrator (Clontech) following the manufacturer’s instructions. The concentrated virus was stored in aliquots at −80 °C.