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"Reducing double-stranded RNA formation in in vitro transcription using a novel RNA polymerase"

A robust and versatile enzyme enabling seamless integration into RNA synthesis protocols

Double-stranded RNA (dsRNA) is a significant contaminant in mRNA. During in vitro transcription (IVT), RNA polymerases naturally generate dsRNA byproducts through a number of mechanisms including cryptic promoters or loopback RNAs. Once inside cells or animals, these dsRNA contaminants can then trigger innate immune activation and diminish protein expression, thereby jeopardizing the safety, tolerability, and efficacy of the potential therapeutic product.

Recently, novel mutant enzymes have been developed to replace the commonly used wild-type T7 RNA polymerase to minimize dsRNA formation in IVT. While a theoretically simple approach, some enzymes may necessitate IVT condition optimization — especially when used in co-transcriptional capping — to avoid adverse effects on mRNA quality attributes such as yield, integrity, and capping efficiency.

In this tech note, we characterized a robust and versatile RNA polymerase engineered for seamless integration into your existing RNA synthesis protocols. Unlike some alternatives that may require tedious optimization, this enzyme minimizes dsRNA formation across various constructs and capping methods without sacrificing yield, integrity, or capping efficiency.

 

From this tech note, you'll learn how this novel RNA polymerase:

• Dramatically reduces dsRNA across diverse RNA constructs with or without base modifications
• Effortlessly substitutes standard T7 RNA polymerase in IVT reactions
• Enables efficient synthesis of long and complex sequences like saRNAs with minimal dsRNA
• Leads to improved mRNA performance: lower inflammatory response, higher protein expression

 

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