The aim of our project is to contribute a robust reporter system as well as a stable RNA polymerase expression system.
Our project focuses on two primary objectives:
The development of our novel reporter system for T7 RNA polymerase allows for the identification of different variants in the downstream experiments. Furthermore, the reporter system can be further tweaked by replacing the promoter sequence to characterize other RNA polymerases, or by extending the transcript length to assess the transcriptional strength of these RNA polymerases. Our reporter system allows for the identification of RNA polymerase variants without the use of antibiotics, a safer alternative to previously established studies.
The development of T7 RNAP involves employing techniques such as site-directed mutagenesis, ancestral reconstruction, and random mutagenesis. The aim of these methods is to create optimized variants of T7 RNAP that show improved transcriptional efficiency and stability, making them suitable for a wider range of biotechnological applications, including the synthesis of modified RNAs for therapeutic use.
By integrating our results from both aspects of the project, the generated T7 RNAP variants can be tested within the reporter system to assess their performance, ultimately leading to a versatile platform that can be adapted for various applications with further modifications. This dual approach not only enhances the understanding of T7 RNAP’s capabilities but also provides a valuable tool for future research in gene expression and synthetic biology.