ArticleNucleic acids research2017
Twin-primer non-enzymatic DNA assembly: an efficient and accurate multi-part DNA assembly method.
Article in Nucleic acids research, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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Who cites it
23 citing papers in PubMed.
- High-throughput synthesis of DNA fragments by molecular self-assembly of overlapping oligonucleotides.Nature biotechnology · 2026Article
- Advanced applications of synthetic biology technology in biosynthesis of bioactive compounds from medicinal plants.Chinese herbal medicines · 2026Review
- DAPE cloning with modified primers for producing designated lengths of 3' single-stranded ends in PCR products.PloS one · 2025Article
- Advancements in Golden Gate Cloning: A Comprehensive Review.Methods in molecular biology (Clifton, N.J.) · 2025Review
- Neq2X7: a multi-purpose and open-source fusion DNA polymerase for advanced DNA engineering and diagnostics PCR.BMC biotechnology · 2024Article
- PlasmidMaker is a versatile, automated, and high throughput end-to-end platform for plasmid construction.Nature communications · 2022Article
- Recombinase-free cloning (RFC) protocol for gene swapping.Molecular biology research communications · 2022Article
- Recent Advances in Strategies for the Cloning of Natural Product Biosynthetic Gene Clusters.Frontiers in bioengineering and biotechnology · 2021Review
- Exploring the synthetic biology potential of bacteriophages for engineering non-model bacteria.Nature communications · 2020Review
- Pantoea sp. P37 as a novel nonpathogenic host for the heterologous production of rhamnolipids.MicrobiologyOpen · 2020Article
- An artificial chromosome ylAC enables efficient assembly of multiple genes in Yarrowia lipolytica for biomanufacturing.Communications biology · 2020Article
- Metagenomics Approaches in Discovery and Development of New Bioactive Compounds from Marine Actinomycetes.Current microbiology · 2020Review
- Genetic platforms for heterologous expression of microbial natural products.Natural product reports · 2019Review
- COMPASS for rapid combinatorial optimization of biochemical pathways based on artificial transcription factors.Nature communications · 2019Article
- Standardizing Automated DNA Assembly: Best Practices, Metrics, and Protocols Using Robots.SLAS technology · 2019Article
- T5 exonuclease-dependent assembly offers a low-cost method for efficient cloning and site-directed mutagenesis.Nucleic acids research · 2019Article
- Start-Stop Assembly: a functionally scarless DNA assembly system optimized for metabolic engineering.Nucleic acids research · 2019Article
- A seamless and iterative DNA assembly method named PS-Brick and its assisted metabolic engineering for threonine and 1-propanol production.Biotechnology for biofuels · 2019Article
- Direct cloning and heterologous expression of natural product biosynthetic gene clusters by transformation-associated recombination.Methods in enzymology · 2019Article
- Nimble Cloning: A Simple, Versatile, and Efficient System for Standardized Molecular Cloning.Frontiers in bioengineering and biotechnology · 2019Article
Corrections and comments
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Authors and funding
5 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
DNA assembly forms the cornerstone of modern synthetic biology. Despite the numerous available methods, scarless multi-fragment assembly of large plasmids remains challenging. Furthermore, the upcoming wave in molecular biological automation demands a rethinking of how we perform DNA assembly. To streamline automation workflow and minimize operator intervention, a non-enzymatic assembly method is highly desirable. Here, we report the optimization and operationalization of a process called Twin-Primer Assembly (TPA), which is a method to assemble polymerase chain reaction-amplified fragments into a plasmid without the use of enzymes. TPA is capable of assembling a 7 kb plasmid from 10 fragments at ∼80% fidelity and a 31 kb plasmid from five fragments at ∼50% fidelity. TPA cloning is scarless and sequence independent. Even without the use of enzymes, the performance of TPA is on par with some of the best in vitro assembly methods currently available. TPA should be an invaluable addition to a synthetic biologist's toolbox.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.