Co-engineering DNAzymes and Phagemid Systems for Efficient and Scalable Production of Functional Single-Stranded DNA Nanostructures
6 affiliations
- 1East China University of Science and Technology , , ,
- 2Shanghai Institute of Applied Physics, Chinese Academy of Sciences , , ,
- 3Fudan University , , ,
- 4Shanghai Jiao Tong University , , ,
- 5Shanghai University , , ,
- 6Jiangxi Normal University , , ,
Abstract
DNA nanostructures have been widely explored for diverse applications, yet their scalable and cost-efficient production remains a major limitation. Phagemid-based biosynthesis of single-stranded DNA (ssDNA) provides a promising approach, but its efficiency is constrained by limited deoxyribozyme (DNAzyme) performance and suboptimal system design. Here, we develop an optimized bioproduction platform for ssDNA nanostructures by simultaneously engineering DNAzymes and phagemid-based production systems. Sequence-guided DNAzyme design identifies determinants for catalytic efficiency and pH adaptability, enabling highly efficient and robust in cis cleavage. The optimized DNAzymes achieve 100% self-cleavage within 30 min and expand the operational pH range from a narrow window around 7.2 to 6.6–7.8, thus enabling more reliable production of target ssDNA for structural folding. By combining optimization of phagemid constructs and helper phage/plasmid combinations, the resulting platform supports the biotechnological production of different nanostructures, including square and aptamer-functionalized triangle assemblies. With the p3024 phagemid and VCSM13 helper phage, production of functionalized triangular ssDNA nanostructures was successfully scaled to a 5 L bioreactor, yielding 151.0 mg/L phagemid ssDNA. Doxorubicin-loaded, aptamer AS1411-functionalized triangular nanostructures (DOX@DN-AS1411) enabled targeted and rapid intracellular drug delivery in breast cancer cells. This work establishes a scalable platform to produce programmable DNA nanostructures, facilitating their translation toward biomedical and nanomanufacturing applications.
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BibTeX
@article{hu2026coengineering,
title = {{Co-engineering DNAzymes and Phagemid Systems for Efficient and Scalable Production of Functional Single-Stranded DNA Nanostructures}},
author = {Jinyi Hu and Wanqing Ding and Ning Jiang and Yujie Wang and Junjun Hou and Hongzhou Gu and Linjie Guo and Lihua Wang and Jiang Li and Jiaofang Huang and Jiangchao Qian},
journal = {JACS Au},
year = {2026},
month = nov,
doi = {10.1021/jacsau.6c01405},
url = {https://pubs.acs.org/jaaucr/article/doi/10.1021/jacsau.6c01405/5462646/Co-engineering-DNAzymes-and-Phagemid-Systems-for},
note = {CC BY-NC-ND 4.0},
}
RIS
TY - JOUR
TI - Co-engineering DNAzymes and Phagemid Systems for Efficient and Scalable Production of Functional Single-Stranded DNA Nanostructures
AU - Jinyi Hu
AU - Wanqing Ding
AU - Ning Jiang
AU - Yujie Wang
AU - Junjun Hou
AU - Hongzhou Gu
AU - Linjie Guo
AU - Lihua Wang
AU - Jiang Li
AU - Jiaofang Huang
AU - Jiangchao Qian
PY - 2026
DA - 2026/11/03
JO - JACS Au
DO - 10.1021/jacsau.6c01405
UR - https://pubs.acs.org/jaaucr/article/doi/10.1021/jacsau.6c01405/5462646/Co-engineering-DNAzymes-and-Phagemid-Systems-for
AB - DNA nanostructures have been widely explored for diverse applications, yet their scalable and cost-efficient production remains a major limitation. Phagemid-based biosynthesis of single-stranded DNA (ssDNA) provides a promising approach, but its efficiency is constrained by limited deoxyribozyme (DNAzyme) performance and suboptimal system design. Here, we develop an optimized bioproduction platform for ssDNA nanostructures by simultaneously engineering DNAzymes and phagemid-based production systems. Sequence-guided DNAzyme design identifies determinants for catalytic efficiency and pH adaptability, enabling highly efficient and robust in cis cleavage. The optimized DNAzymes achieve 100% self-cleavage within 30 min and expand the operational pH range from a narrow window around 7.2 to 6.6–7.8, thus enabling more reliable production of target ssDNA for structural folding. By combining optimization of phagemid constructs and helper phage/plasmid combinations, the resulting platform supports the biotechnological production of different nanostructures, including square and aptamer-functionalized triangle assemblies. With the p3024 phagemid and VCSM13 helper phage, production of functionalized triangular ssDNA nanostructures was successfully scaled to a 5 L bioreactor, yielding 151.0 mg/L phagemid ssDNA. Doxorubicin-loaded, aptamer AS1411-functionalized triangular nanostructures (DOX@DN-AS1411) enabled targeted and rapid intracellular drug delivery in breast cancer cells. This work establishes a scalable platform to produce programmable DNA nanostructures, facilitating their translation toward biomedical and nanomanufacturing applications.
SN - 2691-3704
ER -