(naked DNA) articles in PubMed
- Detection of parvovirus B19 DNA in blood: Viruses or DNA remnants? [Journal Article]
- J Clin Virol 2016 Sep 13; 84:19-23JC
- CONCLUSIONS: Detection of B19V DNA in blood by PCR does not necessarily imply that B19V replication takes place and that infectious B19V virions are present. We propose that remnant B19V DNA strands can be released from tissues without active replication. This finding urges to reconsider an assumed role of B19V infection mainly based on B19V DNA detection in blood, a much debated subject in clinical syndromes such as myocarditis and arthritis.
- Multiplexed instrument-free meningitis diagnosis on a polymer/paper hybrid microfluidic biochip. [Journal Article]
- Biosens Bioelectron 2016 Sep 12; 87:865-873BB
- Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae), and Haemophilus influenzae type b (Hib) are three most common pathogens accounting for most bacterial meningitis, a...
Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae), and Haemophilus influenzae type b (Hib) are three most common pathogens accounting for most bacterial meningitis, a serious global infectious disease with high fatality, especially in developing nations. Because the treatment and antibiotics differ among each type, the identification of the exact bacteria causing the disease is vital. Herein, we report a polymer/paper hybrid microfluidic biochip integrated with loop-mediated isothermal amplification (LAMP) for multiplexed instrument-free diagnosis of these three major types of bacterial meningitis, with high sensitivity and specificity. Results can be visually observed by the naked eye or imaged by a smartphone camera under a portable UV light source. Without using any specialized laboratory instrument, the limits of detection of a few DNA copies per LAMP zone for N. meningitidis, S. pneumoniae and Hib were achieved within 1h. In addition, these three types of microorganisms spiked in artificial cerebrospinal fluid (ACSF) were directly detected simultaneously, avoiding cumbersome sample preparation procedures in conventional methods. Compared with the paper-free non-hybrid microfluidic biochip over a period of three months, the hybrid microfluidic biochip was found to have a much longer shelf life. Hence, this rapid, instrument-free and highly sensitive microfluidic approach has great potential for point-of-care (POC) diagnosis of multiple infectious diseases simultaneously, especially in resource-limited settings.
- Direct Oxidative Damage of Naked DNA Generated upon Absorption of UV Radiation by Nucleobases. [Journal Article]
- J Phys Chem Lett 2016 Sep 19JP
- It has been shown that in addition to formation of pyrimidine dimers, UV irradiation of DNA in the absence of photosensitizer also induces formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine, but the me...
It has been shown that in addition to formation of pyrimidine dimers, UV irradiation of DNA in the absence of photosensitizer also induces formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine, but the mechanism of formation of that oxidized base has not been clearly established. In the present study, we provide an unambiguous demonstration that absorption of UVC and UVB radiation by the nucleobases induces DNA oxidation via a direct process (one-electron oxidation) and not singlet oxygen. Evidence arose from the fact that polyamine-guanine adducts that are specifically produced through the transient formation of guanine radical cation are generated following UV irradiation of DNA in presence of polyamine even in the absence of any photosensitizer.
- Four Novel Splice-Switch Reporter Cell Lines: Distinct Impact of Oligonucleotide Chemistry and Delivery Vector on Biological Activity. [Journal Article]
- Nucleic Acid Ther 2016 Sep 15NA
- New advances in oligonucleotide (ON) chemistry emerge continuously, and over the last few years, several aspects of ON delivery have been improved. However, clear knowledge regarding how certain chem...
New advances in oligonucleotide (ON) chemistry emerge continuously, and over the last few years, several aspects of ON delivery have been improved. However, clear knowledge regarding how certain chemistries behave alone, or in combination with various delivery vectors, is limited. Moreover, characterization is frequently limited to a single reporter cell line and, when different cell types are studied, experiments are commonly not carried out under similar conditions, hampering comparative analysis. To address this, we have developed a small "tissue" library of new, stable, pLuc/705 splice-switching reporter cell lines (named HuH7_705, U-2 OS_705, C2C12_705, and Neuro-2a_705). Our data show that, indeed, the cell type used in activity screenings influences the efficiency of ONs of different chemistry (phosphorothioate with locked nucleic acid or 2'-O-methyl with or without N,N-diethyl-4-(4-nitronaphthalen-1-ylazo)-phenylamine). Likewise, the delivery method, Lipofectamine(®) 2000, PepFect14 nanoparticles, or "naked" uptake, also demonstrates cell-type-dependent outcomes. Taken together, these cell lines can potentially become useful tools for future in vitro evaluation of new nucleic acid-based oligomers as well as delivery compounds for splice-switching approaches and cell-specific therapies.
- Establishment of an on-site diagnostic procedure for detection of orf virus from oral lesions of Japanese serows (Capricornis crispus) by loop-mediated isothermal amplification. [Journal Article]
- J Vet Med Sci 2016 Sep 15JV
- Orf virus infection has been prevalent continuously in the population of wild Japanese serows (Capricornis crispus), goat-like grazing cloven-hoofed mammal species that live mainly in mountainous are...
Orf virus infection has been prevalent continuously in the population of wild Japanese serows (Capricornis crispus), goat-like grazing cloven-hoofed mammal species that live mainly in mountainous areas of Japan. Currently, definitive diagnosis of infection requires time-consuming laboratory work. To diagnose rapidly on-site, we developed a field-friendly procedure for the detection of orf virus from oral cavity lesions. DNA was extracted from goat saliva spiked with orf virus as a proxy for Japanese serows by a commercial kit without the use of electricity, and the quality of the extracted DNA was evaluated by conventional polymerase chain reaction (PCR). Extracted DNA was amenable to DNA amplification, the same as when extracted in a laboratory. Next, to find optimal conditions for DNA amplification by loop-mediated isothermal amplification (LAMP), Bst and Csa DNA polymerases and 3 colorimetric indicators for visual diagnosis, hydroxy naphthol blue (HNB), malachite green and D-QUICK, were compared using a portable cordless incubator. The combination of Bst or Csa DNA polymerase with HNB was found to be easiest for visual diagnosis by the naked eye, and viral DNA was successfully amplified from all orf virus strains used. These results suggest that the procedure established here can work completely on-site and can be useful for definitive diagnosis and differentiation of orf virus infection in Japanese serows in remote mountainous areas.
- Rapid and Sensitive Detection of Didymella bryoniae by Visual Loop-Mediated Isothermal Amplification Assay. [Journal Article]
- Front Microbiol 2016; 7:1372FM
- Didymella bryoniae is a pathogenic fungus that causes gummy stem blight (GSB) in Cucurbitaceae crops (e.g., cantaloupe, muskmelon, cucumber, and watermelon). GSB produces lesions on the stems and lea...
Didymella bryoniae is a pathogenic fungus that causes gummy stem blight (GSB) in Cucurbitaceae crops (e.g., cantaloupe, muskmelon, cucumber, and watermelon). GSB produces lesions on the stems and leaves, and can also be spread by seeds. Here, we developed a rapid, visual, and sensitive loop-mediated amplification (LAMP) assay for D. bryoniae detection based on sequence-characterized amplified regions (GenBank accession nos GQ872461 and GQ872462) common to the two random amplification of polymorphic DNA group genotypes (RGI and RGII) of D. bryoniae; ideal conditions for detection were optimized for completion in 45 min at 63°C. The sensitivity and specificity of the LAMP assay were further analyzed in comparison with those of a conventional polymerase chain reaction (PCR). The sensitivity of the LAMP assay was 1000-fold higher than that of conventional PCR with a detection limit of 0.1 fg μL(-1) of targeted DNA. The LAMP assay could be accomplished in about 45 min, with the results visible to the naked eye. The assay showed high specificity in discriminating all D. bryoniae isolates from seven other fungal pathogens that occur in Cucurbitaceae crops. The LAMP assay also detected D. bryoniae infection in young muskmelon leaves with suspected early symptoms of GSB disease. Hence, the technique has great potential for developing rapid and sensitive visual detection methods for the D. bryoniae pathogen in crops and seeds. This method has potential application in early prediction of disease and reducing the risk of epidemics.
- Development and evaluation of real-time loop-mediated isothermal amplification assay for rapid detection of cystic echinococcosis. [Journal Article]
- BMC Vet Res 2016; 12:202BV
- CONCLUSIONS: The developed LAMP assay would be expected to prove highly significant in epidemiological surveys of CE in developing countries or areas of resource-poor settings for both ease of use and cost.
- A simple, fast, label-free colorimetric method for detection of telomerase activity in urine by using hemin-graphene conjugates. [Journal Article]
- Biosens Bioelectron 2016 Sep 2; 87:600-606BB
- Telomerase, a widely accepted cancer biomarker for early cancer diagnostics, is considered as an important therapeutic target. To now, it is still a challenging subject to develop a simple and sensit...
Telomerase, a widely accepted cancer biomarker for early cancer diagnostics, is considered as an important therapeutic target. To now, it is still a challenging subject to develop a simple and sensitive strategy for telomerase activity detection. Herein, we reported a simple colorimetric strategy for label-free quantification of human telomerase activity in urine by using hemin-graphene nanomaterial (H-GNs). H-GNs possessed tailored dispersibility in the high salt concentration and highly active biomimetic oxidation catalyst property. In this strategy, H-GNs were adjusted to coagulate to appropriate degree by carefully selecting the contained NaCl amount in the presence of original TS primer. The supernatant of the solution contained few H-GNs and showed light blue color. Under the action of telomerase, TS primer was elongated with repeating sequences of (TTAGGG)n. These negatively charged DNA enhanced individual H-GNs electrostatic repulsion and resisted salt-induced H-GNs coagulation. As a result, the supernate of the corresponding solution containing more dispersed H-GNs and showed dark blue color after chromogenic reaction. Thus, telomerase activity could be quasi-quantified by naked eye and precise quantified by UV spectrometer. The proposed method has the linear range from 100 to 2300 HeLa cells/mL and the detection limit was 60 cells/mL. It has been successfully applied to detect telomerase activity in real urine samples. Obtained results were in good agreement with the clinical diagnosis. Therefore, this colorimetric approach affords simplicity, sensitivity and reliability in telomerase activity detection.
- Synthesis and Characterization of Cell-Permeable Oligonucleotides Bearing Reduction-Activated Protecting Groups on the Internucleotide Linkages. [Journal Article]
- Bioconjug Chem 2016 Sep 21; 27(9):2149-56BC
- Cell-permeable oligodeoxyribonucleotides (ODNs) bearing reduction-activated protecting groups were synthesized as oligonucleotide pro-drugs. Although these oligonucleotides were amenable to solid-pha...
Cell-permeable oligodeoxyribonucleotides (ODNs) bearing reduction-activated protecting groups were synthesized as oligonucleotide pro-drugs. Although these oligonucleotides were amenable to solid-phase DNA synthesis and purification, the protecting group on their phosphodiester moiety could be readily cleaved by nitroreductase and NADH. Moreover, these compounds exhibited good nuclease resistance against 3'-exonuclease and endonuclease and good stability in human serum. Fluorescein-labeled ODNs modified with reduction-activated protecting groups showed better cellular uptake compared with that of naked ODNs.
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- Comparative study of three magnetic nano-particles (FeSO4, FeSO4/SiO2, FeSO4/SiO2/TiO2) in plasmid DNA extraction. [Journal Article]
- Anal Biochem 2016 Nov 15; 513:68-76AB
- Recent updates on Magnetic Nano-Particles (MNPs) based separation of nucleic acids have received more attention due to their easy manipulation, simplicity, ease of automation and cost-effectiveness. ...
Recent updates on Magnetic Nano-Particles (MNPs) based separation of nucleic acids have received more attention due to their easy manipulation, simplicity, ease of automation and cost-effectiveness. It has been indicated that DNA molecules absorb on solid surfaces via hydrogen-bonding, and hydrophobic and electrostatic interactions. These properties highly depend on the surface condition of the solid support. Therefore, surface modification of MNPs may enhance their functionality and specification. In the present study, we functionalized Fe3O4 nano-particle surface utilizing SiO2 and TiO2 layer as Fe3O4/SiO2 and Fe3O4/SiO2/TiO2 and then compare their functionality in the adsorption of plasmid DNA molecules with the naked Fe3O4 nano-particles. The result obtained showed that the purity and amount of DNA extracted by Fe3O4 coated by SiO2 or SiO2/TiO2 were higher than the naked Fe3O4 nano-particles. Furthermore, we obtained pH 8 and 1.5 M NaCl as an optimal condition for desorption of DNA from MNPs. The result further showed that, 0.2 mg nano-particle and 10 min at 55 °C are the optimal conditions for DNA desorption from nano-particles. In conclusion, we recommended Fe3O4/SiO2/TiO2 as a new MNP for separation of DNA molecules from biological sources.