AffiGEN ® T7 RNA Polymerase GMP Grade
AffiGEN® RNA Polymerase is a recombinant, GMP-grade mutant of bacteriophage T7 RNA polymerase (200 U/µL) expressed in E. coli (clone D05). It specifically recognizes T7 promoters and synthesizes RNA from ssDNA or dsDNA templates, with reduced dsRNA by-products and enhanced capping efficiency. All production steps avoid animal-derived materials and ampicillin, ensuring suitability for mRNA vaccine applications.
Product Specifications
| Component | Description |
| Enzyme | T7 RNA Polymerase (200 U/µL) |
| Buffer | 10× Transcription Buffer, GMP grade |
| Available Sizes | 1 mL, 5 mL, 20 mL |
| Activity | 200 U/µL (1 U incorporates 1 nmol [³H] ATP in 1 h at 37 °C, pH 8.0) |
| Optimal Temp | 37°C |
| Storage | Store at -20±5°C; transport at ≤0°C |
Quality Standards
| Test | Specification | Method |
| Purity | ≥ 95% | HPLC |
| Enzyme Activity | ≥ 200 U/µL | Fluorescence assay |
| DNA Nucleases | ≤10% degradation | Incubation with pUC19 |
| RNase | Negative | Fluorescence assay |
| Exogenous DNA | < 120 fg/µL | qPCR |
| Host Proteins | < 50 ppm | Protein residue assay |
| Endotoxin | <2 EU/mL | Photometric assay |
Applications Notes
- In vitro transcription of mRNA, siRNA, gRNA (10 – 14 000 nt).
- One-step capping protocols with CAG trimer cap analog ("AG" initiation > 90% capping).
- Recommended enzyme concentration: 5 – 30 U/µL in a 20 µL reaction at 37 °C for 2 h.
- Limit enzyme volume to ≤ 20% of total reaction; max 7 freeze-thaw cycles.
- Use high-purity, RNase-free DNA (OD260/280 = 1.8 – 2.0).
- Quantify RNA by fluorescence assay when using modified nucleotides; dsRNA by ELISA.