pLenti Plasmids

pLenti plasmids are vectors used primarily for generating lentiviral particles that can transduce a wide variety of mammalian cells, including those that are hard to transfect. They are key tools in gene delivery systems for functional studies, gene therapy, and stable cell line generation.

Key Features

• Lentiviral Components: pLenti plasmids include essential components for lentivirus production, which are derived from the HIV-1 genome. These components are divided into the following regions:
  • Packaging Sequences: Such as the gag, pol, and rev genes (often found in separate packaging plasmids but sometimes included in the vector for specific systems).
  • Envelope Gene: The env gene, often derived from the vesicular stomatitis virus (VSV-G), which facilitates the entry of the virus into target cells.
• Expression Cassette: Contains a promoter and a gene of interest. The promoter drives the expression of the transgene after integration into the host genome. Common promoters used include the CMV promoter or EF1α promoter for robust expression.
• Selectable Markers: Include antibiotic resistance genes such as puromycin, blasticidin, or neomycin. These markers allow for selection of successfully transduced cells and establishment of stable cell lines.
• Long Terminal Repeats (LTRs): Contain sequences necessary for integration into the host genome and transcription of the integrated transgene. Typically, pLenti plasmids have 5' LTR and 3' LTR regions flanking the expression cassette.
• Self-Inactivating (SIN) Design: Many pLenti vectors are designed to be self-inactivating, meaning that the LTRs are modified to prevent transcription of the viral genes after integration. This design reduces the risk of reactivation of viral replication in transduced cells.

Content

• Backbone: Contains the lentiviral packaging sequences, promoter, expression cassette, and LTR regions.
• Transgene: Gene of interest driven by a promoter for expression in mammalian cells.
• Selectable Marker: Gene for antibiotic resistance to select for transduced cells.
• Envelope Gene (Optional): If not using a separate plasmid for the envelope, the plasmid may contain VSV-G or another envelope gene for packaging.

Applications

• Gene Delivery: Efficiently delivers genes into a wide range of mammalian cells, including non-dividing and primary cells.
• Stable Cell Line Creation: Enables stable integration of the transgene into the host genome for long-term expression.
• Functional Studies: Facilitates the study of gene function, gene regulation, and cellular processes in a controlled manner.
• Gene Therapy: Provides a method for delivering therapeutic genes to treat genetic disorders.