Fusing Allosteric Ribozymes with CRISPR-Cas12a for Efficient Diagnostics of Small Molecule Targets.

The CRISPR-Cas systems are adopted as powerful molecular tools for not only genetic manipulation but also point-of-care diagnostics. However, methods to enable diagnostics of non-nucleic-acid targets with these systems are still limited. Herein, by fusing ligand-dependent allosteric ribozymes with CRISPR-Cas12a, a derived CRISPR-Cas system is created for efficient quantitative analysis of non-nucleic-acid targets in 1-2 h.

Expanding DNA Origami Design Freedom with De Novo Synthesized Scaffolds.

The construction of DNA origami nanostructures is heavily dependent on the folding of the scaffold strand, which is typically a single-stranded DNA genome extracted from a bacteriophage (M13). Custom scaffolds can be prepared in a number of methods, but they are not widely accessible to a broad user base in the DNA nanotechnology community. Here, we explored new design and construction possibilities with custom scaffolds prepared in our cost- and time-efficient production pipeline.

Engineering of Multivalent Membrane-Anchored DNA Frameworks for Precise Profiling of Variable Membrane Permeability During Reversible Electroporation.

Electroporation techniques have emerged as attractive tools for intracellular delivery, rendering promising prospects towards clinical therapies. Transient disruption of membrane permeability is the critical process for efficient electroporation-based cargo delivery. However, smart nanotools for precise characterization of transient membrane changes induced by strong electric pulses are extremely limited.

DNAzyme-based faithful probing and pulldown to identify candidate biomarkers of low abundance.

Biomarker discovery is essential for the understanding, diagnosis, targeted therapy and prognosis assessment of malignant diseases. However, it remains a huge challenge due to the lack of sensitive methods to identify disease-specific rare molecules. Here we present MORAC, molecular recognition based on affinity and catalysis, which enables the effective identification of candidate biomarkers with low abundance.

R-loop-dependent promoter-proximal termination ensures genome stability.

The proper regulation of transcription is essential for maintaining genome integrity and executing other downstream cellular functions. Here we identify a stable association between the genome-stability regulator sensor of single-stranded DNA (SOSS) and the transcription regulator Integrator-PP2A (INTAC). Through SSB1-mediated recognition of single-stranded DNA, SOSS-INTAC stimulates promoter-proximal termination of transcription and attenuates R-loops associated with paused RNA polymerase II to prevent R-loop-induced genome instability.

Small DNAs That Specifically and Tightly Bind Transition Metal Ions.

Specific DNA-binding to metal ions is a long-standing fundamental research topic with great potential to transform into nano/biotechnology and therapeutics applications. Herein, based on the mobility change of DNA in denaturing gels, we develop a selection strategy to discover a series of 40-45 nt small DNAs that can bind Zn and Cd specifically and tightly. The Zn- and Cd-bound DNA complexes can even tolerate harsh denaturing conditions of 8 M urea and 50 mM EDTA.

Targeting Tumor Necrosis Factor Receptor 1 with Selected Aptamers for Anti-Inflammatory Activity.

Tumor necrosis factor-α (TNFα) inhibitors are widely used in treating autoimmune diseases like rheumatoid arthritis (RA). These inhibitors can presumably alleviate RA symptoms by blocking TNFα-TNF receptor 1 (TNFR1)-mediated pro-inflammatory signaling pathways. However, the strategy also interrupts the survival and reproduction functions conducted by TNFα-TNFR2 interaction and causes side effects.

A nuclease-mimetic platinum nanozyme induces concurrent DNA platination and oxidative cleavage to overcome cancer drug resistance.

Platinum (Pt) resistance in cancer almost inevitably occurs during clinical Pt-based chemotherapy. The spontaneous nucleotide-excision repair of cancer cells is a representative process that leads to Pt resistance, which involves the local DNA bending to facilitate the recruitment of nucleotide-excision repair proteins and subsequent elimination of Pt-DNA adducts. By exploiting the structural vulnerability of this process, we herein report a nuclease-mimetic Pt nanozyme that can target cancer cell nuclei and induce concurrent DNA platination and oxidative cleavage to overcome Pt drug resistance.

Catalytic DNA-Assisted Mass Production of Arbitrary Single-Stranded DNA.

Synthetic single-stranded (ss) DNA is a cornerstone for life and materials science, yet the purity, quantity, length, and customizability of synthetic DNA are still limiting in various applications. Here, we present PECAN, paired-end cutting assisted by DNAzymes (DNA enzymes or deoxyribozymes), which enables mass production of ssDNA of arbitrary sequence (up to 7000 nucleotides, or nt) with single-base precision. At the core of PECAN technique are two newly identified classes of DNAzymes, each robustly self-hydrolyzing with minimal sequence requirement up- or down-stream of its cleavage site.

Programmable Assembly of Amphiphilic DNA through Controlled Cholesterol Stacking.

The excellent programmability and modifiability of DNA has enabled chemists to reproduce a series of specific molecular interactions in self-assembled synthetic systems. Among diverse modifications, cholesterol conjugation can turn DNA into an amphiphilic molecule (cholesterol-DNA), driving the formation of DNA assemblies through the cholesterol-endowed hydrophobic interaction. However, precise control of such an assembly process remains difficult because of the unbiased accumulation of cholesterol.

Nucleic Acids for Potential Treatment of Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a common systemic inflammatory autoimmune disease that severely affects the life quality of patients. Current therapeutics in clinic mainly focus on alleviating the development of RA or relieving the pain of patients. The emerging biological disease-modifying antirheumatic drugs (DMARDs) require long-term treatment to achieve the expected efficacy.

DNA nanotechnology-facilitated ligand manipulation for targeted therapeutics and diagnostics.

Ligands, mostly binding to proteins to form complexes and catalyze chemical reactions, can serve as drug and probe molecules, as well as sensing elements. DNA nanotechnology can integrate the high editability of DNA nanostructures and the biological activity of ligands into functionalized DNA nanostructures in a manner of controlled ligand stoichiometry, type, and arrangement, which provides significant advantages for targeted therapeutics and diagnostics. As therapeutic agents, multiple- and multivalent-ligands functionalized DNA nanostructures increase ligand-receptor affinity and activate multivalent ligand-receptor interactions, enabling improved regulation of cell signaling and enhanced control of cell behavior.

Small DNAs that Bind Nickel(II) Specifically and Tightly.

Metal recognition by nucleic acids provides an intriguing route for biosensing of metal. Toward this goal, a key prerequisite is the acquisition of nucleic acids that can selectively respond to specific metals. Herein, we report for the first time the discovery of two small DNAs that can specifically bind Ni and discriminate against similar ions, particularly, Co.

Novel approaches of the nanotechnology-based drug delivery systems for knee joint injuries: A review.

The knee joint is one of the largest, most complex, and frequently utilized organs in the body. It is very vulnerable to injuries due to activities, diseases, or accidents, which lead to or cause knee joint injuries in people of all ages. There are several types of knee joint injuries such as contusions, sprains, and strains to the ligament, tendon injuries, cartilage injuries, meniscus injuries, and inflammation of synovial membrane.

New DNA-hydrolyzing DNAs isolated from an ssDNA library carrying a terminal hybridization stem.

DNA-hydrolyzing DNAs represent an attractive type of DNA-processing catalysts distinctive from the protein-based restriction enzymes. The innate DNA property has enabled them to readily join DNA-based manipulations to promote the development of DNA biotechnology. A major in vitro selection strategy to identify these DNA catalysts relies tightly on the isolation of linear DNAs processed from a circular single-stranded (ss) DNA sequence library by self-hydrolysis.

Biotechnological production of ssDNA with DNA-hydrolyzing deoxyribozymes.

Preparation of long single-stranded (ss)DNA in large quantities with high efficiency and purity remains a synthetic challenge. Here, we present a protocol for using DNA-hydrolyzing DNA enzymes (deoxyribozymes) for efficient biotechnological production of milligrams of ssDNA with a customizable sequence up to a few kilobases. Our protocol provides a convenient yet economical way to store the sequence information of target ssDNA on phages for selective mass production on demand.

Engineering Allosteric Ribozymes to Detect Thiamine Pyrophosphate in Whole Blood.

Thiamine deficiency contributes to several human diseases including Alzheimer's. As its biologically active form, thiamine pyrophosphate (TPP) has been considered as a potential biomarker for Alzheimer's disease (AD) based on several clinical reports that apparently lower blood TPP levels were found in patients with mild cognitive impairment to AD. However, highly sensitive and high-throughput detection of TPP in biological fluids remains an analytical challenge.

Programming CircLigase Catalysis for DNA Rings and Topologies.

Circular single-stranded (ss) DNA is an essential element in rolling circle amplification and many DNA nanotechnology constructions. It is commonly synthesized from linear ssDNA by a ligase, which nevertheless suffers from low and inconsistent efficiency due to the simultaneous formation of concatemeric byproducts. Here, we design an intramolecular terminal hybridization strategy to program the ring formation catalytic process of CircLigase, a thermostable RNA ligase 1 that can ligate ssDNA in an intramolecular fashion.

Novel aptasensor-based assay of sonic hedgehog ligand for detection of portal vein invasion of hepatocellular carcinoma.

The high expression of sonic hedgehog ligand (SHh) is closely correlated to the metastasis, drug resistance and poor prognosis of hepatocellular carcinoma (HCC). Therefore, sensitive, specific and efficient detection methods for SHh are needed for the early diagnosis and assessment of prognosis. Herein, an aptamer, AP32 that specifically binds to SHh (K = 25.

A SAM-I riboswitch with the ability to sense and respond to uncharged initiator tRNA.

All known riboswitches use their aptamer to senese one metabolite signal and their expression platform to regulate gene expression. Here, we characterize a SAM-I riboswitch (SAM-I) from the Xanthomonas campestris that regulates methionine synthesis via the met operon. In vitro and in vivo experiments show that SAM-I controls the met operon primarily at the translational level in response to cellular S-adenosylmethionine (SAM) levels.

Engineered DNA nanodrugs alleviate inflammation in inflammatory arthritis.

Rheumatoid arthritis (RA) is an autoimmune disease featured with chronic joint inflammation. Suppression of inflammation is critical to RA treatment and joint protection. In this study, DNA nanodrugs are prepared via the conjugation of NF-κB decoy oligodeoxynucleotides (dODNs) and VCAM-1 targeted peptides (P) onto self-assembled DNA tetrahedrons (TDs).

Near-Atomic Fabrication with Nucleic Acids.

Nucleic acids hold great promise for bottom-up construction of nanostructures programmable self-assembly. Especially, the emerging of advanced sequence design principles and the maturation of chemical synthesis of nucleic acids together have led to the rapid development of structural DNA/RNA nanotechnology. Diverse nucleic acids-based nano objects and patterns have been constructed with near-atomic resolutions and with controllable sizes and geometries.

Targeting nucleolin to obstruct vasculature feeding with an intelligent DNA nanorobot.

A prototype of DNA nanorobot with the ability to transport molecular payloads was designed to target cancer cells in tissue culture. Moreover, a further step was taken to succeed in the first in vivo application of the DNA nanorobot for cancer therapy. The robot was constructed using aptamer and DNA origami to fold a 90-nm tubular device to carry the blood coagulation protease thrombin inside, shielded from circulating platelets and plasma fibrinogen.