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The production of lentiviral vectors centers around the use of a packaging cell to produce the viral vector particles. Large-scale manufacturing of vectors begins with the growth of an adequate number of these packaging cells, such as derivatives of the HEK293T cell line. Generation of vectors for use in the clinical setting requires the use of current Good Manufacturing Practices (cGMP) to ensure the production of high-quality vectors of verified identity, potency, and purity. The steps required for vector production are not complex; however, producing robust, cGMP-compliant production systems for lentiviral vectors has proven a bit more challenging than for producing retroviral vectors. Although lentiviral vector manufacturing has many parallels to gammaretroviral vector production, one of the critical differences that significantly affects the manufacturing process consistency and increases costs of lentiviral vectors is the lack of a packaging cell line with stable transfection of the core packaging plasmids.