eprintid: 10191433
rev_number: 6
eprint_status: archive
userid: 699
dir: disk0/10/19/14/33
datestamp: 2024-04-30 10:26:28
lastmod: 2024-04-30 10:26:28
status_changed: 2024-04-30 10:26:28
type: article
metadata_visibility: show
sword_depositor: 699
creators_name: Sari, Yustika
creators_name: Sousa Rosa, Sara
creators_name: Jeffries, Jack
creators_name: Marques, Marco PC
title: Comprehensive evaluation of T7 promoter for enhanced yield and quality in mRNA production
ispublished: pub
divisions: UCL
divisions: B04
divisions: C05
divisions: F47
keywords: Nucleic-acid therapeutics, RNA, Synthetic biology
note: © 2024 Springer Nature Limited. This article is licensed under a Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/).
abstract: The manufacturing of mRNA vaccines relies on cell-free based systems that are easily scalable and flexible compared with the traditional vaccine manufacturing processes. Typically, standard processes yield 2 to 5 g L−1 of mRNA, with recent process optimisations increasing yields to 12 g L−1. However, increasing yields can lead to an increase in the production of unwanted by-products, namely dsRNA. It is therefore imperative to reduce dsRNA to residual levels in order to avoid intensive purification steps, enabling cost-effective manufacturing processes. In this work, we exploit sequence modifications downstream of the T7 RNA polymerase promoter to increase mRNA yields whilst simultaneously minimising dsRNA. In particular, transcription performance was optimised by modifying the sequence downstream of the T7 promoter with additional AT-rich sequences. We have identified variants that were able to produce higher amounts of mRNA (up to 14 g L−1) in 45 min of reaction. These variants exhibited up to a 30% reduction in dsRNA byproduct levels compared to a wildtype T7 promoter, and have similar EGFP protein expression. The results show that optimising the non-coding regions can have an impact on mRNA production yields and quality, reducing overall manufacturing costs.
date: 2024
date_type: published
publisher: Springer Science and Business Media LLC
official_url: http://dx.doi.org/10.1038/s41598-024-59978-5
oa_status: green
full_text_type: pub
language: eng
primo: open
primo_central: open_green
verified: verified_manual
elements_id: 2271215
doi: 10.1038/s41598-024-59978-5
lyricists_name: Marques, Marco Paulo
lyricists_id: MMARQ87
actors_name: Marques, Marco Paulo
actors_id: MMARQ87
actors_role: owner
full_text_status: public
publication: Scientific Reports
volume: 14
number: 1
article_number: 9655
citation:        Sari, Yustika;    Sousa Rosa, Sara;    Jeffries, Jack;    Marques, Marco PC;      (2024)    Comprehensive evaluation of T7 promoter for enhanced yield and quality in mRNA production.                   Scientific Reports , 14  (1)    , Article 9655.  10.1038/s41598-024-59978-5 <https://doi.org/10.1038/s41598-024-59978-5>.       Green open access   
 
document_url: https://discovery.ucl.ac.uk/id/eprint/10191433/1/s41598-024-59978-5.pdf