Long single-stranded DNA (ssDNA) is a versatile molecular reagent with applications including RNA-guided genome engineering and DNA nanotechnology, yet its production is typically resource-intensive. We introduce a novel method utilizing an engineered Escherichia coli 'helper' strain and phagemid system that simplifies long ssDNA generation to a straightforward transformation and purification procedure. Our method obviates the need for helper plasmids and their associated contamination by integrating M13mp18 genes directly into the E. coli chromosome. We achieved ssDNA lengths ranging from 504 to 20 724 nt with titers up to 250 μg/l following alkaline lysis purification. The efficacy of our system was confirmed through its application in primary T-cell genome modifications and DNA origami folding. The reliability, scalability and ease of our approach promise to unlock new experimental applications requiring large quantities of long ssDNA.
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http://dx.doi.org/10.1093/nar/gkae189 | DOI Listing |
Pathogens
December 2024
Department of Microbiology & Immunology, University of Texas Medical Branch, Galveston, TX 77555, USA.
Flaviviruses, a group of single-stranded RNA viruses spread by mosquitoes or ticks, include several significant neurotropic viruses, such as West Nile virus (WNV), Japanese encephalitis virus (JEV), tick-borne encephalitis virus (TBEV), and Zika virus (ZIKV). These viruses can cause a range of neurological diseases during acute infection, from mild, flu-like symptoms to severe and fatal encephalitis. A total of 20-50% of patients who recovered from acute flavivirus infections experienced long-term cognitive issues.
View Article and Find Full Text PDFJ Phys Chem B
January 2025
College of Chemistry, Beijing Normal University, Beijing 100875, P. R. China.
Under conditions that are close to the real cellular environment, the human telomeric single-stranded overhang (∼200 nt) consisting of tens of TTAGGG repeats tends to form higher order structures of multiple G-quadruplex (G4) blocks. On account of the higher biological relevance of higher order G4 structures, ligand compounds binding to higher order G4 are significant for the drug design toward inhibiting telomerase activity. Here, we study the interaction between a cationic porphyrin derivative, 5,10,15,20-tetra{4-[2-(1-methyl-1-piperidinyl)propoxy]phenyl}porphyrin (T4), and a human telomeric G4-dimer (AG(TAG)) in the mimic intracellular molecularly crowded environment (PEG as a crowding agent) and K or Na solution (i.
View Article and Find Full Text PDFBiosens Bioelectron
January 2025
Education Department of Guangxi Zhuang Autonomous Region, Laboratory of Optic-electric Chemo/Biosensing and Molecular Recognition, Guangxi Collaborative Innovation Center for Chemistry and Engineering of Forest Products, Guangxi Key Laboratory of Chemistry and Engineering of Forest Products, Key Laboratory of Chemistry and Engineering of Forest Products, State Ethnic Affairs Commission, School of Chemistry and Chemical Engineering, Guangxi Minzu University, Nanning 530006, China. Electronic address:
Sugarcane smut is a widespread fungal disease, which severely impairs the quality and sugar yield of sugarcane. Early detection is crucial for mitigating its impact, which makes the development of a highly sensitive and accurate detection method essential. Herein, the Mn-doped zeolite imidazolate framework (ZIF-67), synthesized via a nano-confined-reactor approach, is designed to significantly enhance electron transport and boost the enzyme loading capacity within biofuel cells, thereby potentially enhancing their overall performance.
View Article and Find Full Text PDFSmall
January 2025
Institute of Molecular Medicine and Shanghai Key Laboratory for Nucleic Acid Chemistry and Nanomedicine, State Key Laboratory of Oncogenes and Related Genes, Department of Laboratory Medicine, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, China.
A 3D DNA spatial chip (DSC) based on an icosahedral DNA origami framework is introduced to construct customized circular single-stranded DNA (c-ssDNA) for data storage. Within the confined space of the DSC, thirty addressable location sequences extending from the framework edges are available for designing circular paths and directing the assembly of a series of information oligonucleotides for efficient ligation. This strategy is verified by constructing c-ssDNAs from up to 15 fragments to encode two poems (800 and 860 nucleotides).
View Article and Find Full Text PDFNat Commun
January 2025
Developmental Therapeutics Branch & Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, USA.
Type IA topoisomerases (TopoIAs) are present in all living organisms. They resolve DNA/RNA catenanes, knots and supercoils by breaking and rejoining single-stranded DNA/RNA segments and allowing the passage of another nucleic acid segment through the break. Topoisomerase III-β (TOP3B), the only RNA topoisomerase in metazoans, promotes R-loop disassembly and translation of mRNAs.
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