- The in vitro T7 transcription system is a widely used tool in molecular biology that harnesses the powerful and specific activity of T7 RNA polymerase to synthesize RNA transcripts from DNA templates in a controlled, cell-free environment.
- This system relies on a linear or circular DNA template containing a T7 promoter upstream of the sequence to be transcribed.
- When purified T7 RNA polymerase, ribonucleotide triphosphates (NTPs), and the appropriate buffer components are added, the polymerase initiates transcription at the T7 promoter and synthesizes RNA with high efficiency and fidelity.
- Because T7 RNA polymerase does not require additional transcription factors and has minimal sequence requirements beyond the promoter, the system is remarkably simple, robust, and scalable.
- In vitro T7 transcription is extensively used to generate RNA probes, mRNA for translation studies, synthetic guide RNAs (sgRNAs) for CRISPR/Cas9 systems, and RNA standards for qPCR or diagnostics.
- It is also fundamental to mRNA vaccine production, where large quantities of capped and polyadenylated RNA are needed.
- Variants of this system include co-transcriptional capping strategies and modified nucleotides to enhance RNA stability or translational efficiency.
- Its cell-free nature offers advantages such as the absence of RNases from living cells, freedom from regulatory interference, and suitability for high-purity or modified RNA production.
- Overall, the in vitro T7 transcription system is a versatile and indispensable platform for RNA synthesis across basic research, biotechnology, and therapeutics.
