Dual UTR-A novel 5′ untranslated region design for synthetic biology applications
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Abstract
Bacterial 5′ untranslated regions of mRNA (UTR) involve in a complex regulation of gene expression; however, the exact sequence features contributing to gene regulation are not yet fully understood. In this study, we report the design of a novel 5′ UTR, dual UTR, utilizing the transcriptional and translational characteristics of 5′ UTRs in a single expression cassette. The dual UTR consists of two 5′ UTRs, each separately leading to either increase in transcription or translation of the reporter, that are separated by a spacer region, enabling de novo translation initiation. We rationally create dual UTRs with a wide range of expression profiles and demonstrate the functionality of the novel design concept in Escherichia coli and Pseudomonas putida using different promoter systems and coding sequences. Overall, we demonstrate the application potential of dual UTR design concept in various synthetic biology applications ranging from fine-tuning of gene expression to maximization of protein production.
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The 5′ UTR is mainly involved in translation of its downstream ORF sequence, and the Kozak sequence is generally added after the 5′ UTR sequence to improve translation efficiency.
“The 5′ UTR is mainly involved in translation of its downstream ORF sequence.132,133 The Kozak sequence is generally added after the 5′ UTR sequence to improve translation efficiency.134”
From Advances in COVID-19 mRNA vaccine development (2022) · cites this paper as [132]
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Cites
Molecular Cloning: A Laboratory Manual
Sambrook, J., Fritsch, E., & Maniatis, T. (1989) · UCLA - Biblioteca de Ciencias de la Salud · Cited by 133,589 · see citing works
Molecular cloning: A laboratory manual
Deininger, P. L. (1990) · Analytical Biochemistry · doi:10.1016/0003-2697(90)90595-z · Cited by 84,684 · see citing works
Automated design of synthetic ribosome binding sites to control protein expression
Salis, H. M., Mirsky, E. A., & Voigt, C. A. (2009) · Nature Biotechnology · doi:10.1038/nbt.1568 · Cited by 1,844 · see citing works
Coding-Sequence Determinants of Gene Expression in <i>Escherichia coli</i>
Kudla, G., Murray, A. W., Tollervey, D., & Plotkin, J. B. (2009) · Science · doi:10.1126/science.1170160 · Cited by 1,458 · see citing works
Gene regulation by riboswitches
Mandal, M., & Breaker, R. R. (2004) · Nature Reviews Molecular Cell Biology · doi:10.1038/nrm1403 · Cited by 913 · see citing works
Precise and reliable gene expression via standard transcription and translation initiation elements
Mutalik, V. K., Guimaraes, J. C., Cambray, G., Lam, C. S. C., Christoffersen, M. J., Mai, Q.-A., Tran, A. B., Paull, M. L., Keasling, J. D., Arkin, A. P., et al. (2013) · Nature Methods · doi:10.1038/nmeth.2404 · Cited by 748 · see citing works
Regulation of translation via mRNA structure in prokaryotes and eukaryotes
Kozak, M. (2005) · Gene · doi:10.1016/j.gene.2005.06.037 · Cited by 747 · see citing works
STEITZ, J. A., & Jakes, K. S. (1975) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.72.12.4734 · Cited by 692 · see citing works
[4] Plasmid transformation of Escherichia coli and other bacteria
Hanahan, D., Jessee, J. A., & Bloom, F. R. (1991) · Methods in enzymology on CD-ROM/Methods in enzymology · doi:10.1016/0076-6879(91)04006-a · Cited by 621 · see citing works
Cooperation Between Translating Ribosomes and RNA Polymerase in Transcription Elongation
Proshkin, S., Rahmouni, A. R., Mironov, A. S., & Nudler, E. (2010) · Science · doi:10.1126/science.1184939 · Cited by 565 · see citing works
Combinatorial engineering of intergenic regions in operons tunes expression of multiple genes
Pfleger, B. F., Pitera, D. J., Smolke, C. D., & Keasling, J. D. (2006) · Nature Biotechnology · doi:10.1038/nbt1226 · Cited by 535 · see citing works
Nikel, P. I., & de Lorenzo, V. (2018) · Metabolic Engineering · doi:10.1016/j.ymben.2018.05.005 · Cited by 469 · see citing works
Cardinale, S., & Arkin, A. P. (2012) · Biotechnology Journal · doi:10.1002/biot.201200085 · Cited by 450 · see citing works
Codon influence on protein expression in E. coli correlates with mRNA levels
Boël, G., Letso, R. R., Neely, H., Price, W. N., Wong, K., Su, M., Luff, J. D., Valecha, M., Everett, J. K., Acton, T., et al. (2016) · Nature · doi:10.1038/nature16509 · Cited by 429 · see citing works
Kosuri, S., Goodman, D. B., Cambray, G., Mutalik, V. K., Gao, Y., Arkin, A. P., Endy, D., & Church, G. M. (2013) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1301301110 · Cited by 392 · see citing works
Seo, S. W., Yang, J.-S., Kim, I., Yang, J., Min, B. E., Kim, S., & Jung, G. Y. (2013) · Metabolic Engineering · doi:10.1016/j.ymben.2012.10.006 · Cited by 300 · see citing works
Cambray, G., Guimaraes, J. C., & Arkin, A. P. (2018) · Nature Biotechnology · doi:10.1038/nbt.4238 · Cited by 235 · see citing works
Control of Prokaryotic Translational Initiation by mRNA Secondary Structure
de Smit, M. H., & DUIN, J. V. (1990) · Progress in nucleic acid research and molecular biology · doi:10.1016/s0079-6603(08)60707-2 · Cited by 216 · see citing works
RNA processing enables predictable programming of gene expression
Qi, L. S., Haurwitz, R. E., Shao, W., Doudna, J. A., & Arkin, A. P. (2012) · Nature Biotechnology · doi:10.1038/nbt.2355 · Cited by 198 · see citing works
Small regulatory non‐coding RNAs in bacteria: physiology and mechanistic aspects
Repoila, F., & Darfeuille, F. (2009) · Biology of the Cell · doi:10.1042/bc20070137 · Cited by 179 · see citing works
Blatny, J. M., Brautaset, T., Winther-Larsen, H. C., Karunakaran, P., & Valla, S. (1997) · Plasmid · doi:10.1006/plas.1997.1294 · Cited by 171 · see citing works
Interdependence of translation, transcription and mRNA degradation in the lacZ gene
Yarchuk, O., Jacques, N., Guillerez, J., & Dreyfus, M. (1992) · Journal of Molecular Biology · doi:10.1016/0022-2836(92)90617-s · Cited by 170 · see citing works
The intricate world of riboswitches
Coppins, R. L., Hall, K. B., & Groisman, E. A. (2007) · Current Opinion in Microbiology · doi:10.1016/j.mib.2007.03.006 · Cited by 139 · see citing works
Comparison of mRNA features affecting translation initiation and reinitiation
Osterman, I. А., Evfratov, S. A., Владимирович, С. П., & Dontsova, O. А. (2012) · Nucleic Acids Research · doi:10.1093/nar/gks989 · Cited by 135 · see citing works
Balzer, S., Pettersen, V. K., Megerle, J. A., Lale, R. R., Brautaset, T., & Valla, S. (2013) · Microbial Cell Factories · doi:10.1186/1475-2859-12-26 · Cited by 129 · see citing works
Self-citationDirect Detection of Abortive RNA Transcripts in Vivo
Goldman, S. R., Ebright, R. H., & Nickels, B. E. (2009) · Science · doi:10.1126/science.1169237 · Cited by 124 · see citing works
Levin-Karp, A., Barenholz, U., Bareia, T., Dayagi, M., Zelcbuch, L., Antonovsky, N., Noor, Ε., & Milo, R. (2013) · ACS Synthetic Biology · doi:10.1021/sb400002n · Cited by 122 · see citing works
Craig, M. L., Tsodikov, O. V., McQuade, K. L., Schlax, P. E., Capp, M. W., Saecker, R. M., & Record, M. T. (1998) · Journal of Molecular Biology · doi:10.1006/jmbi.1998.2129 · Cited by 114 · see citing works
Nouaille, S., Mondeil, S., Finoux, A.-L., Moulis, C., Girbal, L., & Cocaign‐Bousquet, M. (2017) · Nucleic Acids Research · doi:10.1093/nar/gkx781 · Cited by 87 · see citing works
Na, D., Lee, S., & Lee, D. (2010) · BMC Systems Biology · doi:10.1186/1752-0509-4-71 · Cited by 85 · see citing works
Multiple activities of RNA-binding proteins S1 and Hfq
Hajnsdorf, E., & Boni, I. V. (2012) · Biochimie · doi:10.1016/j.biochi.2012.02.010 · Cited by 81 · see citing works
Bakke, I., Berg, L., Aune, T. E. V., Brautaset, T., Sletta, H., Tøndervik, A., & Valla, S. (2009) · Applied and Environmental Microbiology · doi:10.1128/aem.02315-08 · Cited by 55 · see citing works
Initial Transcribed Sequence Mutations Specifically Affect Promoter Escape Properties
Hsu, L. M., Cobb, I. M., Ozmore, J. R., Khoo, M., Nahm, G., Xia, L., Bao, Y., & Ahn, C. (2006) · Biochemistry · doi:10.1021/bi060247u · Cited by 53 · see citing works
Berg, L., Lale, R. R., Bakke, I., Burroughs, N. J., & Valla, S. (2009) · Microbial Biotechnology · doi:10.1111/j.1751-7915.2009.00107.x · Cited by 44 · see citing works
Self-citationPm Promoter Expression Mutants and Their Use in Broad-Host-Range RK2 Plasmid Vectors
Winther-Larsen, H. C., Blatny, J. M., Valand, B., Brautaset, T., & Valla, S. (2000) · Metabolic Engineering · doi:10.1006/mben.1999.0143 · Cited by 32 · see citing works
Lale, R. R., Berg, L., Stüttgen, F., Netzer, R., Stafsnes, M. H., Brautaset, T., Aune, T. E. V., & Valla, S. (2011) · Applied and Environmental Microbiology · doi:10.1128/aem.02091-10 · Cited by 28 · see citing works
Self-citationHeggeset, T. M. B., Pettersen, V. K., Nærdal, I., Valla, S., Sletta, H., Ellingsen, T. E., & Brautaset, T. (2012) · Applied and Environmental Microbiology · doi:10.1128/aem.02407-12 · Cited by 25 · see citing works
Broad-Host-Range Plasmid Vectors for Gene Expression in Bacteria
Lale, R. R., Brautaset, T., & Valla, S. (2011) · Methods in molecular biology · doi:10.1007/978-1-61779-197-0_19 · Cited by 18 · see citing works
Self-citationPettersen, V. K., Strand, T. A., Almaas, E., Naas, A. E., Brautaset, T., & Valla, S. (2013) · PLoS ONE · doi:10.1371/journal.pone.0066429 · Cited by 6 · see citing works
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Fang, E., Liu, X., Li, M., Zhang, Z., Song, L., Zhu, B., Wu, X., Liu, J., Zhao, D., & Li, Y. (2022) · Signal Transduction and Targeted Therapy · doi:10.1038/s41392-022-00950-y · Cited by 601 · see citing works
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Pourseif, M. M., Masoudi‐Sobhanzadeh, Y., Azari, E., Parvizpour, S., Barar, J., Ansari, R. A., & Omidi, Y. (2022) · Drug Discovery Today · doi:10.1016/j.drudis.2022.103341 · Cited by 54 · see citing works
Zhang, J., Liu, Y., Li, C., Xiao, Q., Zhang, D., Chen, Y., Rosenecker, J., Ding, X., & Guan, S. (2023) · Pharmaceutics · doi:10.3390/pharmaceutics15092182 · Cited by 39 · see citing works
Deal, C., Wannemaeker, L. D., & Mey, M. D. (2024) · FEMS Microbiology Reviews · doi:10.1093/femsre/fuae004 · Cited by 29 · see citing works
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Tietze, L., & Lale, R. R. (2021) · Microbial Biotechnology · doi:10.1111/1751-7915.13868 · Cited by 29 · see citing works
Self-citationSahajpal, N., Lai, C.-Y. J., Hastie, A. R., Mondal, A. K., Dehkordi, S. R., van der Made, C. I., Fedrigo, O. D., Al-Ajli, F. O., Jalnapurkar, S., Byrska‐Bishop, M. S., et al. (2022) · iScience · doi:10.1016/j.isci.2022.103760 · Cited by 21 · see citing works
Bartolo-Aguilar, Y., Chávez-Cabrera, C., Flores‐Cotera, L. B., Badillo-Corona, J. A., Oliver-Salvador, C., & Marsch, R. (2022) · Journal of Genetic Engineering and Biotechnology · doi:10.1186/s43141-022-00455-9 · Cited by 21 · see citing works
Immethun, C. M., Kathol, M., Changa, T., & Saha, R. (2022) · Frontiers in Bioengineering and Biotechnology · doi:10.3389/fbioe.2022.800734 · Cited by 17 · see citing works
Tan, Y., Liang, J., Lai, M., Wan, S., Luo, X., & Li, F. (2023) · Biotechnology Advances · doi:10.1016/j.biotechadv.2023.108272 · Cited by 16 · see citing works
Li, H., Yao, D., Ying, J., Han, X., Zhang, X., Fang, X., Fang, Z., & Xiao, Y. (2022) · Biochemical Engineering Journal · doi:10.1016/j.bej.2022.108718 · Cited by 16 · see citing works
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Wong, C. F. A., van Vliet, L., Bhujbal, S. V., Guo, C., Sletmoen, M., Stokke, B. T., Hollfelder, F., & Lale, R. R. (2021) · ACS Synthetic Biology · doi:10.1021/acssynbio.1c00084 · Cited by 13 · see citing works
Self-citationZhang, Y., Bailey, T. S., Kubiak, A. M., Lambin, P., & Theys, J. (2022) · ACS Synthetic Biology · doi:10.1021/acssynbio.2c00401 · Cited by 12 · see citing works
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Zhang, Z., Du, J., Zhang, D., Han, R., Wu, X., & Liang, Y. (2025) · European journal of medical research · doi:10.1186/s40001-025-03060-x · Cited by 16 · see citing works
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Wang, S., Xu, X., Viitala, T., Shu, Y., & Zhang, H. (2025) · Molecular Therapy — Nucleic Acids · doi:10.1016/j.omtn.2025.102737 · Cited by 11 · see citing works
Feng, J., Wang, Q., Qin, Z., Guo, X., Fu, H., Yang, S., & Wang, J. (2024) · Biotechnology and Bioengineering · doi:10.1002/bit.28701 · Cited by 11 · see citing works
Ren, J., Oh, S., & Na, D. (2025) · The Journal of Microbiology · doi:10.71150/jm.2501033 · Cited by 8 · see citing works
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Sahajpal, N., Dean, J. H., Hilton, B., Fee, T., Skinner, C. D., Hastie, A. R., DuPont, B. R., Chaubey, A., Friez, M. J., & Stevenson, R. E. (2025) · Genome Research · doi:10.1101/gr.279318.124 · Cited by 5 · see citing works
Zhou, X., Cao, W., Huang, C., Zhang, X., Zhou, S., & Deng, Y. (2025) · Biotechnology Advances · doi:10.1016/j.biotechadv.2025.108781 · Cited by 4 · see citing works
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Dahiya, G. S., Bakken, T. I., Fages‐Lartaud, M., & Lale, R. R. (2023) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.1101/2023.10.15.562386 · Cited by 1 · see citing works
Self-citation
- Authors
- Balzer Le, S., Onsager, I., Lorentzen, J. A., & Lale, R.
- Affiliations
- PhotoSynLab, Department of Biotechnology, Faculty of Natural Sciences, Norwegian University of Science and Technology, N-7491, Trondheim, Norway
- Norwegian University of Science and Technology (NO)
- Corresponding author
- Onsager, Ingerid
- Lorentzen, Jon Andreas
- Lale, Rahmi
- ORCID
- Lale, Rahmi
- Venue
- Synthetic Biology
- Publisher
- Oxford University Press (OUP)
- Published
- 2020-01-01
- Volume
- 5
- Issue
- 1
- Pages
- ysaa006-ysaa006
- ISSN
- 2397-7000 (ISSN-L)
- PMC
- PMC7476403
- OpenAlex
- W3034698986
- Cited by
- 22
- References
- 39
- Citations by year
- 2025: 5 · 2024: 3 · 2023: 5 · 2022: 7 · 2021: 2
- FWCI
- 1.14
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Synthetic Biology (publishedVersion, cc-by-nc)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
- PubMed Central (publishedVersion, cc-by-nc)
- PubMed (publishedVersion)
- NORA - Norwegian Open Research Archives (submittedVersion, cc-by-nc)
- Funders
- Norwegian University of Science and Technology, Trondheim, Norway
- SINTEF Industry
- Norwegian Research Council (192432, 192123)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Keywords
- 5' untranslated region, mRNA secondary structure, gene expression regulation, transcriptional regulation, translational regulation, spacer region, synthetic promoters, Escherichia coli, Pseudomonas putida, protein production
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