Complementary DNA Chain Research Articles

Novel inner filter effect-based near-infrared electrochemiluminescence sensor mediated by well-matched AgBr-nitrogen-doped Ti3C2 MXene and nonmetallic plasmon WO3•H2O

The development of innovative and efficient strategy is of paramount importance for near-infrared (NIR) electrochemiluminescence (ECL) sensing, which can substantially promote ECL detection in a wide range of situations. Herein, the inner filter effect (IFE) strategy was designed to construct an ultrasensitive NIR ECL biosensor based on the well-matched AgBr nanocrystals (NCs) decorated nitrogen-doped Ti3C2 MXene nanocomposites (AgBr–N–Ti3C2) and hydrated defective tungsten oxide nanosheets (dWO3•H2O). Specifically, the AgBr–N–Ti3C2 nanocomposites displayed extremely effective NIR ECL emission because N-doping could accelerate electron transfer and boost the red-shift of the ECL spectrum. The nonmetallic plasmon dWO3•H2O was used as an absorber due to its facile tuning of the spectra overlap and higher molar extinction coefficients. Time-resolved emission decay curves proved that the decreased ECL intensity was ascribed to the IFE-based steady quenching mechanism. With the support of tetracycline (TC) aptamer and the complementary DNA chain, the fabricated NIR ECL-IFE biosensor performed a wide linear range of 100 nM ∼ 10 fM with a low detection limit of 2.2 fM (S/N = 3), and it exhibited excellent stability, sensitivity, and reproducibility, so as to be applied to real samples. This strategy opens a new avenue to constructing an efficient NIR ECL-IFE system and shows excellent practical potential in actual sample analysis.

Polymorphism of <i>Sdr</i> genes regulating seed dormancy in <i>Triticum persicum</i> Vav. and <i>Triticum aethiopicum</i> Jakubz.

Preharvest sprouting of wheat grain, sporadically observed in many regions of cultivation of this crop, leads to deterioration of its food and sowing qualities. Seed dormancy is considered to be the main component of resistance to preharvest sprouting. This physiological state of seeds is regulated by many genes, and it depends heavily on environmental conditions. One of the regulators of seed dormancy in cereals is the Sdr4 gene (Seed dormancy 4), which was first studied in rice. In common wheat, the homologues of this gene (TaSdr-A1 and TaSdr-B1) are also involved in the regulation of seed dormancy. The search for valuable alleles in local varieties and endemic forms is a promising area of research aimed at increasing the resistance of crops to adverse environmental factors. In this study, Sdr genes were sequenced in several accessions of two tetraploid wheat species with limited cultivation areas: Persian wheat (Triticum persicum Vav.) and Ethiopian wheat (Triticum aethiopicum Jakubz.). As a result, the same Sdr-A1 and Sdr-B1 variants that had been found in common wheat were detected in these species. The Persian wheat accessions possessed only the Sdr-A1a allele, while Ethiopian ones, only Sdr-A1b. The analysis of F2 hybrids obtained from crossing these tetraploid species showed that the Sdr-A1b allele was associated with a lower germination index of grains than Sdr-A1a. This result was inconsistent with earlier association studies. Previously unknown polymorphisms were found in the promoter of the Sdr-B1 gene in the studied accessions. A deletion of 16 nucleotides was detected in the 3’-terminal region of the TraesCS2B02G215200 gene, located on the complementary DNA chain close to the 3’-end of the Sdr-B1 gene. Possible effects of the detected polymorphisms on the expression of Sdr genes are discussed.

Use of β Radiation to Localize the Binding Sites of Mercury Ions and Platinum-Containing Ligand in DNA

Composite sequence-specific ligands with DNA-damaging groups may dramatically increase the efficacy of radiotherapy. The most promising damage sensitizers are the atoms of heavy elements, in which electrons are emitted from upper orbitals and a multiply charged positive ion forms when an electron is kicked out from lower orbitals. The biophysical mechanisms of DNA damage produced by these sensitizers are far from fully understood. In this work, high-performance polyacrylamide gel electrophoresis (PAGE) in denaturing gel was used to investigate the nature of DNA cleavage on exposure to β radiation for complexes of heavy atom-containing ligands with DNA restriction fragments. It was demonstrated for the first time that DNA in complexes with Pt-bis-netropsin or mercury salts is cleaved in the vicinity of the heavy atom in the presence of radioactive isotopes emitting β particles of different energies. In the presence of 1 M glycerol, the cleavage of the DNA sugar-phosphate backbone was almost entirely due to a neutralization of the multiply charged Auger ion and was not associated with the Auger electron electron-beam radiolysis. Based on the observations, a relatively simple technique was proposed for precise localization of binding sites for various DNA ligands containing a heavy atom. Analysis of the end groups at the cleavage point and the nature of damage to the complementary DNA chain made it possible to speculate about the mechanisms of direct influence of irradiation on a heavy atom.

Aptamer-Based Fluorescence Assay for Detection of Isocarbophos and Profenofos

Abstract A fluorescence method for detection of isocarbophos and profenofos was established based on the specific recognition of aptamer. The aptamer (F-ssDNA) recognizing isocarbophos and profenofos was labeled with FAM at 5′-end, and when it bound to complementary DNA chain (Q-ssDNA) labeled with DABCYL at 3′-end, the double stranded structure would be formed and the fluorescence signal would become very weak due to the effect of fluorescent resonance energy transfer. However, the complementary DNA would dissociate from the double-stranded structure when the aptamer recognized the added target molecules, and the fluorescence signal would thus be recovered and become stronger. Based on this principle, isocarbophos and profenofos could be quantitatively detected. The optimized experimental conditions were as follows: 25 nM F-ssDNA and 50 nM Q-ssDNA-2 were incubated for 20 min at 25°C, then the equal volume pesticides were added and incubated for 60 min before the fluorescence intensity of the system was determined. Under the optimal conditions, the change of the fluorescence intensity of the system (ΔI) showed a linear relationship with the isocarbophos or profenofos concentration in the range of 50—500 μM. The detection limit (LOD, 3σ) was 11.4 μM for isocarbophos with the relative standard deviation (RSD) of 5.8% (n = 10). The detection limit was 14.0 μM for profenofos with RSD of 4.9% (n = 10). The recovery ranged from 85.8% to 95.3% when the method was applied to measure isocarbophos and profenofos in real water samples.

An Aptamer-based Fluorescence Assay for Ochratoxin A

Abstract A novel analytical method for the detection of Ochratoxin A (OTA) is established based on the aptamer recognition and fluorescent probe technology. The present method is developed based on the fact that when the immobilized aptamer bonds to the target OTA, it can induce the conformation change of aptamer and result in the dissociation of the FAM-tagged complementary DNA chain from aptamer, finally leading to the fluorescent signal change. Based on it, OTA can be quantified. All the condition factors affecting the performance of the present method are investigated. The results show if the avidin concentration coated on the microplate is 25 mg L−1, the aptamer concentration is 50 nM, the FAM-tagged complementary DNA chain concentration is 150 nM, the binding buffer solution is chosen as 10 mM HEPES, pH 7.0 (contains 120 mM NaCl, 5 mM KCl, 20 mM MgCl2, and 20 mM CaCl2), and the binding reaction is conducted at 45°C for 40 min, the optimal analytical performance can be achieved for the present work. Under the optimal conditions, the linear range for the OTA concentration detection is 2.0 × 10−8-1.0 × 10−5 g L−1 with a detection limit of 1 × 10−8 g L−1. The RSD is 2.6% for 11 parallel measurements of 1 × 10−6 g L−1 OTA. Meanwhile, the present method is highly selective for OTA and easy to be operated. It has been successfully applied to measure OTA content in real samples.

Target Genes of Neuron-Restrictive Silencer Factor Are Abnormally Up-Regulated in Human Myotilinopathy

Myotilinopathy is a subgroup of myofibrillar myopathies caused by mutations in the myotilin gene in which there is aggregation of abnormal cytoskeletal proteins and ubiquitin. We report here on the accumulation of neuron-related proteins such as ubiquitin carboxy-terminal hydrolase L1 (UCHL1), synaptosomal-associated protein 25, synaptophysin, and alpha-internexin in aberrant protein aggregates in myotilinopathy. We have determined that the neuron-restrictive silencer factor (NRSF)/RE1 silencing transcription factor (REST), a transcription factor expressed in non-neuronal tissues repressing the expression of several neuronal genes, is reduced in myotilinopathies. Moreover, NRSF transfection reduces UCHL1, synaptosomal-associated protein 25, synaptophysin, and alpha-internexin mRNA levels in DMS53 cells, whereas short interferring NRSF transfection increases UCHL1 and synaptophysin mRNA levels in U87-MG cells. Chromatin immunoprecipitation assays have shown that NRSF interacts with the UCHL1 promoter in U87-MG and HeLa cells. In silico analysis of the UCHL1 gene promoter sequence using the MatInspector software has predicted three potential neuron-restrictive silencer elements (NRSEs): NRSE1 located in the complementary DNA chain and NRSE2 and NRSE3 in intron 1, in the coding and complementary chains, respectively. Together, these findings show, for the first time, abnormal regulation of NRSF/REST as a mechanism associated with the aberrant expression of selected neuron-related proteins, which in turn accumulate in abnormal protein aggregates, in myotilinopathy.

Adsorption and hybridisation of DNA-surfactants at fluid surfaces and lipid bilayers

In this paper, we explore the properties of a novel class of functional surfactants based on DNA. These DNA-based surfactants are short chain DNA oligonucleotides covalently attached to a large hydrophobic group, which makes the modified DNA molecules amphiphilic and allows DNA chains to be adsorbed and oriented at fluid surfaces. We show that these materials are surface-active at various fluid surfaces, including air–water and oil–water interfaces, as well as lipid bilayers. We demonstrate that once adsorbed the DNA-surfactants used remain on the liquid surface upon hybridisation with a complementary DNA chain. We use complementary DNA-surfactants to functionalise fluid surfaces and to program the interactions between them based on Watson–Crick pairing. By selecting the appropriate DNA base sequences the interaction between the fluid surfaces functionalised with DNA-surfactants can be programmed with the level of specificity of the enzyme–substrate interaction. We also suggest a novel type of aqueous ink based on DNA-functionalised small liposomes for micropatterning of solid surfaces with DNA by a microcontact printing technique. We illustrate the capabilities of this technique by specific deposition of complementary DNA-functionalised liposomes onto DNA-micropatterned solid surfaces.

Properties and Applications of Novel DNA-Based Surfactants

ABSTRACTWe have designed a novel class of functional surfactants based on DNA and have explored their properties in relation to several possible applications. The DNA-surfactants consist of short chain DNA oligonucleotides covalently bound to a large hydrophobic group, which makes the DNA molecules amphiphilic. We demonstrate that these materials behave like common detergents and are surface-active at various fluid surfaces, e.g. air-water, oil-water interfaces, and lipid bilayers. We also show that once adsorbed the DNA-surfactants remain on the liquid surface upon hybridisation with a complementary DNA chain. We use complementary DNA-surfactants to functionalise fluid surfaces and to program the interactions between them based on Watson-Crick pairing. By selecting the appropriate DNA base sequences, the interaction between the fluid surfaces functionalised with DNA-surfactants can be programmed with a very high specificity. We have also developed a novel procedure for micro-patterning of solid surfaces with DNA by a microcontact printing with aqueous inks of DNA-surfactants which can be utilised for rapid fabrication of DNA assays and genetic biochips.

Synthesis of iron-binding oligonucleotides and their reactions with single-stranded DNA

Two 12-base oligonucleotides were synthesized and modified by attaching metal-binding ligands (2,6-pyridinedicarboxylate and 2,2′-dipicolylamine) to their 5′-ends. When hybridized to a 135-base single stranded DNA, in the presence of Fe 2+, O 2, and a reducing agent, these obligonucleotides cleaved the complementary DNA chain with unique and high base-sequence specificities.

The RNA primer synthesized by primase to initiate phage G4 DNA replication.

With phage G4 DNA as template, primase (the dnaG protein) synthesizes a 26- to 29-residue RNA transcript at the origin of replication. The sequence starts with ATP and contains a hairpin region of one A-U and seven G-C base pairs. Covalent linkage between the RNA and the newly synthesized complementary DNA chain indicates that the RNA transcript serves as a primer.