- Home
- IVD
- By Technology Types
- By Diseases Types
- By Product Types
- Research
- Resource
- Distributors
- Company
| Product Name | Lentiviral Packaging Plasmid Mix, Third-Generation System |
| Catalog No. | ATR-CTR-0029 |
| Description | Pre-optimized lentiviral packaging plasmid mixture for third-generation lentivirus production in HEK293T cells. The third-generation (4-plasmid) system separates the HIV-1 gag/pol, rev, and envelope (VSV-G) genes onto three separate packaging plasmids, co-transfected with the transfer vector carrying the transgene of interest. This design minimizes the risk of generating replication-competent lentivirus (RCL) by requiring four recombination events, and eliminates the Tat gene through a chimeric 5' LTR with constitutive CMV or RSV promoter. The premixed packaging plasmids at optimized molar ratios eliminate the need for individual plasmid titration, reducing hands-on time and improving batch-to-batch lentiviral titer consistency. |
| Intended Use | Production of high-titer, replication-incompetent lentiviral particles for stable gene delivery, gene knockout (CRISPR), gene knockdown (shRNA), cDNA overexpression, and library screening in dividing and non-dividing mammalian cells. |
| Principle / Technology | Third-generation 4-plasmid lentiviral system: Transfer vector (contains transgene + packaging signal Ψ + LTRs), pMDLg/pRRE (Gag/Pol), pRSV-Rev (Rev), pMD2.G (VSV-G envelope); co-transfection into HEK293T by calcium phosphate or PEI; self-inactivating (SIN) 3' LTR for improved biosafety. |
| Detection Method | Co-transfect HEK293T cells with transfer vector + packaging plasmid mix using transfection reagent; harvest supernatant at 48 and 72 hours; concentrate by ultracentrifugation or PEG precipitation; titer by qPCR, p24 ELISA, or functional assay (FACS for fluorescent reporter). |
| Sample Type | HEK293T or HEK293FT producer cells; transduces most dividing and non-dividing mammalian cell types. |
| Performance Range / Specifications | Typical titer: 1×10^6-1×10^8 TU/mL (unconcentrated); after concentration: 1×10^8-1×10^10 TU/mL; biosafety: no RCL detected in standard assays. |
| Sensitivity / LOD | Transduction detectable at MOI 0.1-100 depending on target cell type; lentivirus pseudotyped with VSV-G transduces broad cell range. |
| Specificity | VSV-G pseudotyping enables broad tropism; alternative envelope pseudotypes (Eco, Ampho, GALV, measles) can be used for cell-type specificity by substituting envelope plasmid. |
| Reaction Conditions / Protocol | Transfection of 70-80% confluent HEK293T cells; medium exchange 12-16 hours post-transfection; harvest at 48 hr and 72 hr; filter through 0.45 µm; concentrate if needed. |
| Components / Formulation | pMDLg/pRRE (Gag/Pol) plasmid, pRSV-Rev plasmid, pMD2.G (VSV-G) plasmid, premixed at optimized molar ratio (typically 4:2:1); eGFP transfer vector control plasmid; sterile TE buffer for dilution; protocol. |
| Storage Conditions | Store at -20 °C; avoid repeated freeze-thaw cycles; protect from DNase contamination. |
| Shelf Life | 12 months from date of manufacture at -20 °C. |
| Package Specifications | 1 kit (sufficient for ~5 standard 150 mm dish productions or ~10 T75 flask productions), 5 kits. |
| Product Form | Plasmid DNA in TE buffer at 1 µg/µL (premixed packaging mix) or individual plasmids at 0.5-1 µg/µL; A260/A280 1.8-2.0. |
| Quality Control | Each lot tested for plasmid identity (restriction digest pattern), concentration (spectrophotometry), endotoxin <0.1 EU/µg, sterility, and functional titer (co-transfection with eGFP reporter in HEK293T, titer >1×10^6 TU/mL 48 hr supernatant). |
| Key Features | Third-generation 4-plasmid system for maximum biosafety; pre-optimized packaging plasmid mix; eliminates individual plasmid titration; VSV-G pseudotyped for broad tropism; eGFP reporter control included; validated for high-titer lentivirus production. |
| Purity | Plasmid DNA ≥90% supercoiled; endotoxin <0.1 EU/µg; protein and RNA contamination <1%. |
| Concentration | 1 µg/µL in TE buffer, pH 8.0; packaging plasmid mix at optimized molar ratio (4:2:1 Gag/Pol:Rev:VSV-G). |
| Activity / Unit Definition | Functional titer verified by transduction of HEK293T or HT1080 cells with eGFP reporter; titer >1×10^6 TU/mL in unconcentrated supernatant. |
| Molecular Weight | pMDLg/pRRE: ~12.0 kb; pRSV-Rev: ~4.2 kb; pMD2.G: ~5.8 kb. |
| Source / Origin | Plasmid DNA purified from E. coli DH5α or Stbl3; endotoxin-free purification protocol. |
| pH Range / Optimal pH | TE buffer pH 8.0. |
| Shipping Conditions | Ambient temperature (plasmid DNA is stable); cold pack optional. |
| Expiration Date / Stability | 12 months at -20 °C; plasmid DNA stable at 4 °C for 1 month; avoid repeated freeze-thaw cycles. |
| Regulatory / Compliance | For research use only; BSL-2 containment recommended for lentivirus production and transduction. Lentivirus is a biohazard — follow institutional biosafety guidelines. Not for human therapeutic use. |
| Compatibility | Compatible with second-generation transfer vectors (with intact 5' LTR and Ψ packaging signal). Compatible with calcium phosphate, PEI, and lipid-based transfection reagents for HEK293T co-transfection. Transfer vector must be <10 kb including transgene for efficient packaging. |
| Recommended Buffer System | TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0). |
| Application Notes / Precautions | Thaw plasmid aliquots on ice. Do not vortex plasmid DNA — mix by gentle flicking. For optimal titer: use HEK293T cells at passage <20, 70-80% confluence, and scale transfection reagent proportionally. Collect supernatant at 48 hr (peak titer) and 72 hr (additional ~30% yield). Pool harvests and filter through 0.45 µm PES membrane (NOT nitrocellulose which binds lentivirus). For concentration: ultracentrifugation at 70,000 × g for 2 hr, or PEG-8000 precipitation (10% PEG, 4 °C overnight). Aliquot concentrated virus and store at -80 °C; avoid freeze-thaw. |
| Batch-to-Batch Consistency | Plasmid concentration within ±10%; packaging plasmid ratio verified by analytical digest; functional titer within 2-fold of reference lot. |
For research use only, not for clinical use.
|
There is no product in your cart. |
Copyright © 2026 Alta DiagnoTech. All rights reserved.
