AI Article Synopsis
- Efficient cell-free protein expression faces challenges like template degradation and lower yields compared to live cells, often relying on T7 RNA polymerase.
- A new variant, T7Max, has been developed to significantly boost gene expression in in vitro systems, particularly from linear DNA templates.
- The T7Max promoter is easy to use as it requires no protein modifications, making it compatible with existing T7 RNA polymerase-based protein expression systems.
Article Abstract
Background: Efficient cell-free protein expression from linear DNA templates has remained a challenge primarily due to template degradation. In addition, the yields of transcription in cell-free systems lag behind transcriptional efficiency of live cells. Most commonly used in vitro translation systems utilize T7 RNA polymerase, which is also the enzyme included in many commercial kits.
Results: Here we present characterization of a variant of T7 RNA polymerase promoter that acts to significantly increase the yields of gene expression within in vitro systems. We have demonstrated that T7Max increases the yield of translation in many types of commonly used in vitro protein expression systems. We also demonstrated increased protein expression yields from linear templates, allowing the use of T7Max driven expression from linear templates.
Conclusions: The modified promoter, termed T7Max, recruits standard T7 RNA polymerase, so no protein engineering is needed to take advantage of this method. This technique could be used with any T7 RNA polymerase- based in vitro protein expression system.
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Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9872363 | PMC |
| http://dx.doi.org/10.1186/s13036-023-00323-1 | DOI Listing |
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