Search

Graphical abstract Highlights • (Citrate coated gold nanoparticles + plasmid DNA) are exposed to ultra-soft X-rays. • The radiosensitizing effect of GNPs is verified. • The radiosensitizing effect is inversely proportional to the size of the nanoparticles. • Reactive oxygen species are specifically quenched. • Hydrogen peroxide is converted into OH radicals at the surface of GNPs. Abstract Plasmid DNA in aerated aqueous solution is used as a probe to determine whose of the reactive oxygen species (ROS) generated after absorption of ultra-soft X-rays (USX) take part in biomolecule damage in the presence and in absence of Gold Nano-Particles (GNP) and specific scavengers. Citrate-coated GNPs with core sizes of 6, 10 and 25 nm are synthetized and characterized, especially in terms of plasmon band shift, ζ-potential and hydrodynamic radii (respectively 9, 21 and 30 nm). We confirm the radiosensitizing effect of GNP and show that the SSB number per plasmid increases when, for a same mass of gold element, the core size of the gold nanoparticles decreases. Hydroxyl radicals (OH) are scavenged using the positively-charged 2-amino-2-hydroxymethyl-1,3-propanediol (TRIS) and the neutral dimethyl sulfoxide (DMSO) molecules. Due to both negatively-charged environments of DNA and GNP, at identical scavenging capacity, TRIS is more effective at quenching OH than DMSO. The strong radiosensitizing effect of hydroxyl radicals is confirmed. Methanoate anions are then used to transform OH into hydrogen peroxide; the latter being known to be non-aggressive regarding DNA in the absence of easily oxidable metallic ions (Fenton reactions). Surprisingly, in the presence of GNP, high DNA damage yields are observed even though hydrogen peroxide might not be hold as responsible. Conversely, the radiosensitizing effect of GNP is not observed anymore when H 2 O 2 is scavenged using pyruvate ions. We demonstrate that hydrogen peroxide constitutes quite unexpectedly a hidden stock of OH which are activated at the surface of the GNP by decomposition of H 2 O 2 molecules. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Synthetic DNA lattices and natural DNA thin films doped with Ln3+ are constructed. • he optimum concentration of Ln3+ is estimated by analysis of the topological data. • The significances of optical and electrical characteristics are discussed. • Advantages and applications of Ln3+‒doped DNA nanostructures are addressed. Abstract Deoxyribonucleic acid (DNA) and lanthanide ions (Ln3+) exhibit exceptional optical properties that are applicable to the development of nanoscale devices and sensors. Although DNA nanostructures and Ln3+ ions have been investigated for use in the current state of technology for more than a few decades, researchers have yet to develop DNA and Ln3+ based ultra-violet (UV) photodetectors. Here, we fabricate Ln3+ (such as holmium (Ho3+), praseodymium (Pr3+), and ytterbium (Yb3+))‒doped double crossover (DX)‒DNA lattices through substrate-assisted growth and salmon DNA (SDNA) thin films via a simple drop-casting method on oxygen (O 2) plasma-treated substrates for high performance UV photodetectors. Topological (AFM), optical (UV–vis absorption and FTIR), spectroscopic (XPS), and electrical (I ‒ V and photovoltage) measurements of the DX‒DNA and SDNA thin films doped with various concentrations of Ln3+ ([Ln3+]) are explored. From the AFM analysis, the optimum concentrations of various Ln3+ ([Ln3+] O) are estimated (where the phase transition of Ln3+‒doped DX‒DNA lattices takes place from crystalline to amorphous) as 1.2 mM for Ho3+, 1.5 mM for Pr3+, and 1.5 mM for Yb3+. The binding modes and chemical states are evaluated through optical and spectroscopic analysis. From UV–vis absorption studies, we found that as the [Ln3+] was increased, the absorption intensity decreased up to [Ln3+] O , and increased above [Ln3+] O. The variation in FTIR peak intensities in the nucleobase and phosphate regions, and the changes in XPS peak intensities and peak positions detected in the N 1 s and P 2p core spectra of Ln3+‒doped SDNA thin films clearly indicate that the Ln3+ ions are properly bound between the bases (through chemical intercalation) and to the phosphate backbone (through electrostatic interactions) of the DNA molecules. Finally, the I ‒ V characteristics and time-dependent photovoltage of Ln3+‒doped SDNA thin films are measured both in the dark and under UV LED illuminations (λ LED = 382 nm) at various illumination powers. The photocurrent and photovoltage of Ln3+‒doped SDNA thin films are enhanced up to the [Ln3+] O compared to pristine SDNA due to the charge carriers generated from both SDNA and Ln3+ ions upon the absorption of light. From our observations, the photovoltages as function of illumination power suggest higher responsivities, and the photovoltages as function of time are almost constant which indicates the stability and retention characteristics of the Ln3+‒doped SDNA thin films. Hence, our method which provides an efficient doping of Ln3+ into the SDNA with a simple fabrication process might be useful in the development of high-performance optoelectronic devices and sensors. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • A troponin I biosensor composed of DNA 3WJ on Au nanospike was developed. • The DNA 3WJ was rolled to recognition, EC signal transduction and immobilization. • Au nanospike was prepared to increase the detection limit of troponin I. • This study showed label-free EC detection elements for troponin I detection. Abstract Acute myocardial infarction (AMI) is one of the most serious diseases affecting human beings. In this study, in order to rapidly detect AMI disease, the authors fabricated a label-free electrochemical biosensor composed of a multi-functional DNA structure on Au nanospike (AuNS) with a fabricated Au micro-gap electrode which was incorporated with a PCB chip in order to detect cardiac troponin I (cTnI). As a bioprobe, the DNA 3 way-junction (3WJ) was introduced, because the DNA 3WJ has three arms for embodying the multi-functionality. Each piece of DNA was assembled to simultaneously form the DNA 3WJ for cTnI detection, signal transduction, and immobilization, respectively. The assembled DNA 3WJ structure was confirmed by Native-TBM PAGE. Moreover, in order to increase the electrochemical signal sensitivity, AuNS was prepared. The Au micro-gap array is fabricated with a printed circuit board (PCB) chip in order to control each micro-gap electrode panel selectively so as to detect low volumes of cTnI. Then, the DNA strucuture on pAuNS-modified electrode was prepared using the layer-by-layer (LbL) assembly method. FE-SEM and AFM were used to investigate the modified-surface morphology. The cyclic voltammetry (CV) was measured to confirm the cTnI binding to DNA 3WJ-modified electrode. cTnI was detected in the HEPES solution and human serum, respectively. The LOD result exhibited 1.0 pM in HEPES solution and 1.0 pM in 20% diluted human serum, respectively. In addition, the selectivity test was carried out with various proteins as the control experiment. The present study showed label-free, simple fabrication, and easy-to-tailor detection elements for cTnI. [ABSTRACT FROM AUTHOR]

DNA extraction from environmental samples (environmental DNA; eDNA) for metabarcoding‐based biodiversity studies is gaining popularity as a noninvasive, time‐efficient, and cost‐effective monitoring tool. The potential benefits are promising for marine conservation, as the marine biome is frequently under‐surveyed due to its inaccessibility and the consequent high costs involved. With increasing numbers of eDNA‐related publications have come a wide array of capture and extraction methods. Without visual species confirmation, inconsistent use of laboratory protocols hinders comparability between studies because the efficiency of target DNA isolation may vary. We determined an optimal protocol (capture and extraction) for marine eDNA research based on total DNA yield measurements by comparing commonly employed methods of seawater filtering and DNA isolation. We compared metabarcoding results of both targeted (small taxonomic group with species‐level assignment) and universal (broad taxonomic group with genus/family‐level assignment) approaches obtained from replicates treated with the optimal and a low‐performance capture and extraction protocol to determine the impact of protocol choice and DNA yield on biodiversity detection. Filtration through cellulose‐nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit outperformed other combinations of capture and extraction methods, showing a ninefold improvement in DNA yield over the poorest performing methods. Use of optimized protocols resulted in a significant increase in OTU and species richness for targeted metabarcoding assays. However, changing protocols made little difference to the OTU and taxon richness obtained using universal metabarcoding assays. Our results demonstrate an increased risk of false‐negative species detection for targeted eDNA approaches when protocols with poor DNA isolation efficacy are employed. Appropriate optimization is therefore essential for eDNA monitoring to remain a powerful, efficient, and relatively cheap method for biodiversity assessments. For seawater, we advocate filtration through cellulose‐nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit or phenol‐chloroform‐isoamyl for successful implementation of eDNA multi‐marker metabarcoding surveys. The wide array of capture and extraction methods employed in eDNA research might hinder comparability between studies, because the efficiency of target DNA isolation may vary. In this study, we compared DNA isolation efficiency of multiple protocols, as well as the impact of DNA yield on metabarcoding results. Our results demonstrate an increased risk of false‐negative species detection for targeted eDNA approaches when protocols with poor DNA isolation efficacy are employed. [ABSTRACT FROM AUTHOR]

DNA sequences are fundamental for encoding genetic information. The genetic information may be understood not only from symbolic sequences but also from the hidden signals inside the sequences. The symbolic sequences need to be transformed into numerical sequences so the hidden signals can be revealed by signal processing techniques. All current transformation methods encode DNA sequences into numerical values of the same length. These representations have limitations in the applications of genomic signal compression, encryption, and steganography. We propose a novel integer chaos game representation (inter-CGR or iCGR) of DNA sequences and a lossless encoding method DNA sequences by the iCGR. In the iCGR method, a DNA sequence is represented by the iterated function of the nucleotides and their positions in the sequence. Then the DNA sequence can be uniquely encoded and recovered using three integers from iCGR. One integer is the sequence length and the other two integers represent the accumulated distributions of nucleotides in the sequence. The integer encoding scheme can compress a DNA sequence by 2 bits per nucleotide. The integer representation of DNA sequences provides a prospective tool for sequence analysis and operations. [ABSTRACT FROM AUTHOR]

Halogenated nucleosides, such as 5‐iodo‐2′‐deoxyuridine and 5‐iodo‐2′‐deoxycytidine, are incorporated into the DNA of replicating cells to facilitate DNA single‐strand breaks and intra‐ or interstrand crosslinks upon UV irradiation. In this work, it is shown that the naphthyl‐based organoselenium compounds can mediate the dehalogenation of halogenated pyrimidine‐based nucleosides, such as 5‐X‐2′‐deoxyuridine and 5‐X‐2′‐deoxycytidine (X=Br or I). The rate of deiodination was found to be significantly higher than that of the debromination for both nucleosides. Furthermore, the deiodination of iodo‐cytidines was found to be faster than that of iodo‐uridines. The initial rates of the deiodinations of 5‐iodocytosine and 5‐iodouracil indicated that the nature of the sugar moiety influences the kinetics of the deiodination. For both the nucleobases and nucleosides, the deiodination and debromination reactions follow a halogen‐bond‐mediated and addition/elimination pathway, respectively. DNA photosensitizers: Dehalogenation of halogenated nucleosides, which are potential DNA photosensitizers, by organoselenium compounds is described (see picture). Attachment of 2′‐deoxyribose or ribose sugar to the halogenated nucleobases significantly enhances the rate of deiodination reactions. The Se⋅⋅⋅I halogen bonding plays an important role in the kinetics of the deiodination reactions. [ABSTRACT FROM AUTHOR]

Background: Among the four bases, guanine is the most susceptible to damage from oxidative stress. Replication of DNA containing damaged guanines results in G to T mutations. Therefore, the mutations resulting from oxidative DNA damage are generally expected to predominantly consist of G to T (and C to A when the damaged guanine is not in the reference strand) and result in decreased GC content. However, the opposite pattern was reported 16 years ago in a study of prokaryotic genomes. Although that result has been widely cited and confirmed by nine later studies with similar methods, the omission of the effect of shared ancestry requires a re-examination of the reliability of the results. Results: When aerobic and obligate aerobic prokaryotes were mixed together and anaerobic and obligate anaerobic prokaryotes were mixed together, phylogenetic controlled analyses did not detect significant difference in GC content between aerobic and anaerobic prokaryotes. This result is consistent with two generally neglected studied that had accounted for the phylogenetic relationship. However, when obligate aerobic prokaryotes were compared with aerobic prokaryotes, anaerobic prokaryotes, and obligate anaerobic prokaryotes separately using phylogenetic regression analysis, a significant positive association was observed between aerobiosis and GC content, no matter it was calculated from whole genome sequences or the 4-fold degenerate sites of protein-coding genes. Obligate aerobes have significantly higher GC content than aerobes, anaerobes, and obligate anaerobes. Conclusions: The positive association between aerobiosis and GC content could be attributed to a mutational force resulting from incorporation of damaged deoxyguanosine during DNA replication rather than oxidation of the guanine nucleotides within DNA sequences. Our results indicate a grade in the aerobiosis-associated mutational force, strong in obligate aerobes, moderate in aerobes, weak in anaerobes and obligate anaerobes. [ABSTRACT FROM AUTHOR]

A prospective cohort study (German Clinical Trial Registry, No. 00005273) was performed to determine pre-admission colonization rates, hospital acquisition risk factors, subsequent infection rates and colonization persistence including the respective molecular epidemiology and transmission rates of extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae (EPE). A total of 342 EPEs were isolated from rectal swabs of 1,334 patients on admission, at discharge and 6 months after hospitalization. Inclusion criteria were patients’ age > 18 years, expected length of stays > 48 hours, external referral. The EPEs were characterized by routine microbiological methods, a DNA microarray and ERIC-PCR. EPE colonization was found in 12.7 % of admitted patients, with the highest rate (23.8 %) in patients from nursing homes. During hospitalization, 8.1 % of the patients were de novo EPE colonized, and invasive procedures, antibiotic and antacid therapies were independent risk factors. Only 1/169 patients colonized on admission developed a hospital-acquired EPE infection. Escherichia coli was the predominant EPE (88.9 %), and 92.1% of the ESBL phenotypes could be related to CTX-M variants with CTX-M-1/15 group being most frequent (88.9%). A corresponding β-lactamase could not be identified in five isolates. Hospital-acquired EPE infections in patients colonized before or during hospitalization were rare. The diversity of the EPE strains was much higher than that of the underlying plasmids. In seven patients, transmission of the respective plasmid across different species could be observed indicating that the current strain-based surveillance approaches may underestimate the risk of inter-species transmission of resistance genes. [ABSTRACT FROM AUTHOR]

Background: The detection of Trypanosoma cruzi genetic material in clinical samples is considered an important diagnostic tool for Chagas disease. We have previously demonstrated that PCR using clot samples yields greater sensitivity than either buffy coat or whole blood samples. However, phenol-chloroform DNA extraction from clot samples is difficult and toxic. The objective of the present study was to improve and develop a more sensitive method to recover parasite DNA from clot samples for the diagnosis of Chagas disease. Methodology/Principal findings: A total of 265 match pair samples of whole blood–guanidine (GEB) and clot samples were analyzed; 150 were from Chagas seropositive subjects. DNA was extracted from both whole blood-guanidine samples, using a previously standardized methodology, and from clot samples, using a newly developed methodology based on a combination of the FastPrep technique and the standard method for GEB extraction. A qPCR targeting the nuclear satellite sequences was used to compare the sample source and the extraction method. Of the 150 samples from Chagas positive individuals by serology, 47 samples tested positive by qPCR with DNA extracted by both GEB and clot, but an additional 13 samples tested positive only in DNA extracted from clot. No serology-negative samples resulted positive when tested by qPCR. Conclusions: The new methodology for DNA extraction from clot samples improves the molecular diagnosis of Chagas disease. [ABSTRACT FROM AUTHOR]

Ubiquitous and ever increasing use of mobile phones led to the growing concern about the effects of radiofrequency radiation (RFR) emitted by cell phones on biological systems. The aim of this study is to explore whether long-term RFR exposure at different frequencies affects DNA damage and oxidant-antioxidant parameters in the blood and brain tissue of rats. 28 male Sprague Dawley rats were randomly divided into four equal groups (n = 7). They were identified as Group 1: sham-control, Group 2: 900 MHz, Group 3: 1800 MHz, and Group 4: 2100 MHz. Experimental groups of rats were exposed to RFR 2 h/day for 6 months. The sham-control group of rats was subjected to the same experimental condition but generator was turned off. Specific absorption rates (SARs) at brain with 1 g average were calculated as 0.0845 W/kg, 0.04563 W/kg, and 0.03957, at 900 MHz, 1800 MHz, and 2100 MHz, respectively. Additionally, malondialdehyde (MDA), 8-hydroxydeoxyguanosine (8-OHdG), total antioxidant status (TAS), and total oxidant status (TOS) analyses were conducted in the brain tissue samples. Results of the study showed that DNA damage and oxidative stress indicators were found higher in the RFR exposure groups than in the sham-control group. In conclusion, 900-, 1800-, and 2100-MHz RFR emitted from mobile phones may cause oxidative damage, induce increase in lipid peroxidation, and increase oxidative DNA damage formation in the frontal lobe of the rat brain tissues. Furthermore, 2100-MHz RFR may cause formation of DNA single-strand breaks. [ABSTRACT FROM AUTHOR]

During the 15 years since the discovery of type III human interferons [IFN-λ1(IL-29), IFN-λ2(IL-28A), and IFN-λ3(IL-28B)], numerous biological properties such as anticancer, antiviral, and immunomodulatory activities of this new IFN family have been investigated. Several studies have shown that the encapsulation of pcDNA with PLGA nanoparticles (NPs) protects them against DNase enzyme action and increases the efficiency of gene delivery to the cells. The purpose of this study was to encapsulate pcDNA encoding IFN-λ1 (pIFN-λ1) with a simple and cost-effective method using PLGA NPs. The pIFN-λ1-loaded PLGA NPs were produced by a double-emulsion-solvent evaporation method and characterized in terms of size, size distribution, and zeta potential by DLS and morphologically by SEM and TEM. The bioactivity of NPs was also examined by fluorescent microscopy. The results showed that IFN-λ1 expressed by HEK293T cells could protect HepC-2 cells from the cytopathic effects of EMCV. The NPs were spherical in shape with a mean diameter of 380 ± 3 nm, a zeta potential of -3.3 ± 7.6 mV, an encapsulation efficiency of 75 ± 5%, and a loading capacity of 0.83 ± 0.06. The NPs were also bioactive and easily engulfed by RAW264.7 cells. The pIFN-λ1 could be sustainably released from NPs. Due to the facility and affordability of encapsulation of pIFN-λ1 in the PLGA NPs proposed in this study and the advantages of encapsulation by PLGA, it appeared rational to use pIFN-λ1-loaded NPs instead of naked pIFN-λ1 to determine other unexplained activities of this new cytokine or to use it as an alternative or adjunct to current IFN-α therapy. [ABSTRACT FROM AUTHOR]

"Goji" (Lycium barbarum and Lycium chinense) is a generic name for medical plants with a long historical background in the traditional Chinese medicine. With the emerging trend of "Superfoods" several years ago, Goji berries soon became an established product in European countries and not only are the most popular product of traditional Chinese medicine outside of China but to this day one of the symbols of the entire "Superfood" trend. However, since Goji is an umbrella term for different plant species that are closely related, mislabeling and adulterations (unconsciously or purposely) are possible. We carefully verified the identity of Goji reference plant material based on morphological traits, mainly floral structures of several inflorescences of each individual, in order to create a robust background for the downstream applications that were used on those reference plants and additionally on commercial Goji products. We report morphological and molecular based strategies for the differentiation of Lycium barbarum and Lycium chinense. The two different Goji species vary significantly in seed size, with an almost double average seed area in Lycium chinense compared to Lycium barbarum. Differences could be traced on the molecular level as well; using the psbA-trnH barcoding marker, we detected a single nucleotide substitution that was used to develop an easy one-step differentiation tool based on ARMS (amplification refractory mutation system). Two diagnostic primers used in distinct multiplex PCRs yield a second diagnostic band in a subsequent gel electrophoresis for Lycium barbarum or Lycium chinense , respectively. Our ARMS approach is a strong but simple tool to trace either of the two different Goji species. Both the morphological and the molecular analysis showed that all of the tested commercial Goji products contained fruits of the species Lycium barbarum var. barbarum , leading to the assumption that consumer protection is satisfactory. [ABSTRACT FROM AUTHOR]

Abstract Most of the multiple sclerosis (MS) patients are initially diagnosed with relapsing remitting multiple sclerosis (RRMS). Th17 cells and macrophages have been shown to play critical roles in pathogenesis of MS and initiation of CNS tissue damage. MiR-301a have recently been exposed as an activator of STAT3 in Th17 cells as well as an activator of NF-κB in macrophages by targeting PIAS3 and NKRF correspondingly. However, the possible role of miR-301a in RRMS has not yet been elucidated. Herewith, for the first time, we have studied the expression of miR-301a, NKRF and PIAS3 by quantitative real-time PCR and western blotting method in peripheral blood mononuclear cells (PBMCs) of 71 RRMS patients, including two groups of patients in relapse phase (n = 44) and a group of remitting phase patients (n = 28) in comparison to healthy volunteers (n = 28). In this work, we demonstrate a significant upregulation of miR-301a in relapse phase of MS patients compared to healthy controls and remitting phase patients (P <.05). Our findings also showed a striking decrease of NKRF and PIAS3 expression in relapse phase patients, in contrast to miR-301a and, NF-κB and STAT3 downstream genes (SKA2 and RORc) (P <.05). Subsequently, using luciferase reporter system we confirmed that miR-301a directly targets the mRNA encoding PIAS3 and NKRF proteins. We also showed that miR-301a increasing expression is correlated with down-regulation of PIAS3 and NKRF expression in RRMS patients. Our findings suggest that miR-301a, PIAS3 and NKRF play crucial roles in RRMS and could be considered as promising therapeutic targets. Graphical abstract MiR-301a in a positive feedback loop promotes persistent NF-κB activation; miR-301a gene is located between exons 1 and 2 of SKA2 gene in the human genome which contains an authentic κB site. NF-κB coordinately up-regulates the miR-301a and SKA2. NKRF mRNA, which encodes a repressor of NF-κB transcriptional activity, is targeted by miR-301a. Consequently, miR-301a in a positive feedback loop mediates induction of persistent NF-κB activation. MiR-301a promotes Th17 cells differentiation; Th17 cells are differentiated from human naïve CD4+ T-cells in IL-6/23-STAT3 pathway. PIAS3 is able to suppress activation of STAT3-induced gene RORc by blocking the DNA-binding activity of STAT3. MiR-301a promotes Th17 cells differentiation through negative regulation of PIAS3 expression. Unlabelled Image Highlights • miR-301a, SKA2 and RORc expression levels are up-regulated at relapse phase of RRMS. • Showing an increase expression of miR-301a, NKRF and PIAS3 expression levels are reduced at relapse phase of RRMS. • miR-301a directly interacts and suppresses NKRF and PIAS3 transcript sequences. • miR-301a is a pathogenic factor in RRMS and can modulate inflammatory responses. [ABSTRACT FROM AUTHOR]

The compilation of archaeological and genetic data for ancient European human groups has provided persuasive evidence for a complex series of migrations, population replacements and admixture until the Bronze Age. If the Bronze-to-Iron Age transition has been well documented archaeologically, ancient DNA (aDNA) remains rare for the latter period and does not precisely reflect the genetic diversity of European Celtic groups. In order to document the evolution of European communities, we analysed 45 individuals from the Late Iron Age (La Tène) Urville-Nacqueville necropolis in northwestern France, a region recognized as a major cultural contact zone between groups from both sides of the Channel. The characterization of 37 HVS-I mitochondrial sequences and 40 haplogroups provided the largest maternal gene pool yet recovered for the European Iron Age. First, descriptive analyses allowed us to demonstrate the presence of substantial amounts of steppe-related mitochondrial ancestry in the community, which is consistent with the expansion of Bell Beaker groups bearing an important steppe legacy in northwestern Europe at approximately 2500 BC. Second, maternal genetic affinities highlighted with Bronze Age groups from Great Britain and the Iberian Peninsula regions tends to support the idea that the continuous cultural exchanges documented archaeologically across the Channel and along the Atlantic coast (during and after the Bronze Age period) were accompanied by significant gene flow. Lastly, our results suggest a maternal genetic continuity between Bronze Age and Iron Age groups that would argue in favour of a cultural transition linked to progressive local economic changes rather than to a massive influx of allochthone groups. The palaeogenetic data gathered for the Urville-Nacqueville group constitute an important step in the biological characterization of European Iron age groups. Clearly, more numerous and diachronic aDNA data are needed to fully understand the complex relationship between the cultural and biological evolution of groups from the period. [ABSTRACT FROM AUTHOR]

Background: After transplantation, cell-free DNA derived from the donor organ (ddcfDNA) can be detected in the recipient’s circulation. We aimed to quantify ddcfDNA levels in plasma of kidney transplant recipients thereby investigating the kinetics of this biomarker after transplantation and determining biological variables that influence ddcfDNA kinetics in stable and non-stable patients. Materials and methods: From 107 kidney transplant recipients, plasma samples were collected longitudinally after transplantation (day 1–3 months) within a multicenter set-up. Cell-free DNA from the donor was quantified in plasma as a fraction of the total cell-free DNA by next generation sequencing using a targeted, multiplex PCR-based method for the analysis of single nucleotide polymorphisms. A subgroup of stable renal transplant recipients was identified to determine a ddcfDNA threshold value. Results: In stable transplant recipients, plasma ddcfDNA% decreased to a mean (SD) ddcfDNA% of 0.46% (± 0.21%) which was reached 9.85 (± 5.6) days after transplantation. A ddcfDNA threshold value of 0.88% (mean + 2SD) was determined in kidney transplant recipients. Recipients that did not reach this threshold ddcfDNA value within 10 days after transplantation showed a higher ddcfDNA% on the first day after transplantation and demonstrated a higher individual baseline ddcfDNA%. Conclusion: In conclusion, plasma ddcfDNA fractions decreased exponentially within 10 days after transplantation to a ddcfDNA threshold value of 0.88% or less. To investigate the role of ddcfDNA for rejection monitoring of the graft, future research is needed to determine causes of ddcfDNA% increases above this threshold value. [ABSTRACT FROM AUTHOR]

Rapid identification of DNA oxidative damage sites is of great significance for disease diagnosis. In this work, electric field-regulated click reaction surface-enhanced Raman spectroscopy (e-Click-SERS) was developed aiming at the rapid and specific analysis of furfural, the biomarker of oxidative damage to the 5-carbon site of DNA deoxyribose. In e-Click-SERS, cysteamine-modified porous Ag filaments (cys@p-Ag) were prepared and used as electrodes, amine-aldehyde click reaction sites, and SERS substrates. Cysteamine was controlled as an "end-on" conformation by setting the voltage of cys@p-Ag at -0.1 V, which ensures its activity in participating in the amine-aldehyde click reaction during the detection of furfural. Benefiting from this, the proposed e-Click-SERS method was found to be sensitive, rapid-responding, and interference-resistant in analyzing furfural from plasma. The method detection limits of furfural were 5 ng mL -1 in plasma, and the whole "extraction and detection" procedure was completed within 30 min with satisfactory recovery. Interference from 13 kinds of common plasma metabolites was investigated and found to not interfere with the analysis, according to the exclusive adaptation of the amine-aldehyde click reaction. Notably, the e-Click-SERS technique allows in situ analysis of biological samples, which offers great potential to be a point-of-care testing tool for detecting DNA oxidative damage.

Highlights • Our protocol showed good results in the quantitative and qualitative evaluations. • DNA extracted by the adapted protocol obtained the best purity parameters. • DNA extracted by our protocol proved to be well conserved and suitable for use in WGS. • Cost per sample was much lower than that of similar commercial methods. • Our protocol is an alternative with easy implementation in the laboratory routine. [ABSTRACT FROM AUTHOR]

Background: Next-generation sequencing requires sufficient DNA to be available. If limited, whole-genome amplification is applied to generate additional amounts of DNA. Such amplification often results in many chimeric DNA fragments, in particular artificial palindromic sequences, which limit the usefulness of long sequencing reads. Results: Here, we present Pacasus, a tool for correcting such errors. Two datasets show that it markedly improves read mapping and de novo assembly, yielding results similar to these that would be obtained with non-amplified DNA. Conclusions: With Pacasus long-read technologies become available for sequencing targets with very small amounts of DNA, such as single cells or even single chromosomes. [ABSTRACT FROM AUTHOR]

Background: Count data derived from high-throughput deoxy-ribonucliec acid (DNA) sequencing is frequently used in quantitative molecular assays. Due to properties inherent to the sequencing process, unnormalized count data is compositional, measuring relative and not absolute abundances of the assayed features. This compositional bias confounds inference of absolute abundances. Commonly used count data normalization approaches like library size scaling/rarefaction/subsampling cannot correct for compositional or any other relevant technical bias that is uncorrelated with library size. Results: We demonstrate that existing techniques for estimating compositional bias fail with sparse metagenomic 16S count data and propose an empirical Bayes normalization approach to overcome this problem. In addition, we clarify the assumptions underlying frequently used scaling normalization methods in light of compositional bias, including scaling methods that were not designed directly to address it. Conclusions: Compositional bias, induced by the sequencing machine, confounds inferences of absolute abundances. We present a normalization technique for compositional bias correction in sparse sequencing count data, and demonstrate its improved performance in metagenomic 16s survey data. Based on the distribution of technical bias estimates arising from several publicly available large scale 16s count datasets, we argue that detailed experiments specifically addressing the influence of compositional bias in metagenomics are needed. [ABSTRACT FROM AUTHOR]

Abstract: Theoretical work suggests that sexual conflict should promote the maintenance of genetic diversity by the opposing directions of selection on males and females. If such conflict is pervasive, it could potentially lead to genomic heterogeneity in levels of genetic diversity an idea that so far has not been empirically tested on a genomewide scale. We used large‐scale population genomic and transcriptomic data from the collared flycatcher (Ficedula albicollis) to analyse how sexual conflict, for which we use sex‐biased gene expression as a proxy, relates to genetic variability. Here, we demonstrate that the extent of sex‐biased gene expression of both male‐biased and female‐biased genes is significantly correlated with levels of nucleotide diversity in gene sequences and that this correlation extends to diversity levels also in intergenic DNA and introns. We find signatures of balancing selection in sex‐biased genes but also note that relaxed purifying selection could potentially explain part of the observed patterns. The finding of significant genetic differentiation between males and females for male‐biased (and gonad‐specific) genes indicates ongoing sexual conflict and sex‐specific viability selection, potentially driven by sexual selection. Our results thus indicate that sexual antagonism could potentially be considered as one viable explanation to the long‐standing question in evolutionary biology of how genomes can remain so genetically variable in face of strong natural and sexual selection. [ABSTRACT FROM AUTHOR]

Abstract: DNA double‐strand breaks (DSBs) can be repaired by two major pathways: non‐homologous end‐joining (NHEJ) and homologous recombination (HR). DNA repair pathway choice is governed by the opposing activities of 53BP1, in complex with its effectors RIF1 and REV7, and BRCA1. However, it remains unknown how the 53BP1/RIF1/REV7 complex stimulates NHEJ and restricts HR to the S/G2 phases of the cell cycle. Using a mass spectrometry (MS)‐based approach, we identify 11 high‐confidence REV7 interactors and elucidate the role of SHLD2 (previously annotated as FAM35A and RINN2) as an effector of REV7 in the NHEJ pathway. FAM35A depletion impairs NHEJ‐mediated DNA repair and compromises antibody diversification by class switch recombination (CSR) in B cells. FAM35A accumulates at DSBs in a 53BP1‐, RIF1‐, and REV7‐dependent manner and antagonizes HR by limiting DNA end resection. In fact, FAM35A is part of a larger complex composed of REV7 and SHLD1 (previously annotated as C20orf196 and RINN3), which promotes NHEJ and limits HR. Together, these results establish SHLD2 as a novel effector of REV7 in controlling the decision‐making process during DSB repair. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: The latent reservoir is the main barrier on the road to HIV cure, and clinical approaches towards eradication are often evaluated by their effect on proviral DNA. To ensure inclusiveness and representativeness in HIV cure studies, proviral DNA quantification assays that are able to detect all common circulating HIV clades are urgently needed. Here, three HIV DNA assays targeting three different genomic regions were evaluated for their sensitivity and subtype‐tolerance using digital PCR. Methods: A subtype‐B‐specific assay targeting gag (GAG) and two assays targeting conserved sequences in ltr and pol (LTR and JO) were assessed for their sensitivity and subtype‐tolerance in digital PCR (Bio‐Rad QX200), using a panel of serially diluted subtype reference plasmids as well as a panel of clinical isolates. Both panels represent subtypes A, B, C, D, F, G and circulating recombinant forms (CRFs) AE and AG, which together are responsible for 94% of HIV infections worldwide. Results: HIV subtype was observed to greatly affect HIV DNA quantification results. Robust regression analysis of the serially diluted plasmid panel showed that the GAG assay was only able to linearly quantify subtype B, D and G isolates (4/13 reference plasmids, average R2 = 0.99), whereas LTR and JO were able to quantify all tested isolates (13/13 reference plasmids, respective average R2 = 0.99 and 0.98). In the clinical isolates panel, isolates were considered detectable if all replicates produced a positive result. The GAG assay could detect HIV DNA in four out of five subtype B and one out of two subtype D isolates, whereas the LTR and JO assays detected HIV DNA in all twenty‐nine tested isolates. LTR and JO results were found to be equally precise but more precise than GAG. Conclusions: The results demonstrate the need for a careful validation of proviral reservoir quantification assays prior to investigations into non‐B subtype reservoirs. The LTR and JO assays can sensitively and reliably quantify HIV DNA in a panel that represents the worldwide most prevalent subtypes and CRFs (A, B, C, D, AE, F, G and AG), justifying their application in future trials aimed at global HIV cure. [ABSTRACT FROM AUTHOR]

Background: In the United States, blood donor testing for hepatitis B surface antigen (HBsAg) was initiated in the early 1970s. More recently, testing for antibody to hepatitis B core antigen (anti-HBc) and hepatitis B virus (HBV) DNA have been added. The incidence of hepatitis B has been declining. This study reviews the current status of testing and questions the need for continuation of HBsAg testing.Study Design and Methods: From July 2011 to June 2015, a total of 22.4 million donations were serologically tested for HBsAg and anti-HBc and for HBV-DNA by nucleic acid testing (NAT). All reactive results were evaluated and a subset of donations that were either potential NAT yield (seronegative) or serologically positive but nonreactive by HBV NAT in minipools (MPs) of 16 were further evaluated by individual donation (ID)-NAT. Samples with detectable HBV DNA were defined as actively infected and considered potentially infectious.Results: Routine testing plus supplemental ID-NAT identified 2035 samples representing active infection including 1965 with anti-HBc, 1602 with HBsAg, and 1453 with HBV DNA by MP-NAT, for respective rates per hundred-thousand donations of 9.10, 8.78, 7.16, and 6.50, continuing the downward trend previously observed. There were 29 HBV DNA-yield samples (1:771,389), 35 HBsAg-yield samples (anti-HBc nonreactive), and 404 with occult hepatitis B infection. There were six samples with HBsAg and HBV DNA detectable only by ID-NAT in the absence of anti-HBc; additional testing was consistent with extremely low or negligible levels of DNA.Conclusions: Point estimates of HBV infection rates among blood donors continue to decline, as do those for incidence and residual risk. Elimination of HBsAg screening would have negligible impact, with a risk less than 1 per 4 million donations. [ABSTRACT FROM AUTHOR]

Four benzylidene ketone derivatives were synthesized by reacting 4-nitrobenzaldehyde with several ketone derivatives. The synthesized compounds A, B, C and D had shown antioxidant activity against deoxyribose degradation, while compound C showing the highest activity. The molecular docking simulation indicated that the superior activity of compound C among other compounds can be attributed to the alkyl elongation at ketone chain. [ABSTRACT FROM AUTHOR]

Highlights • We carry out a comparison of likelihood ratios obtained with either a continuous or a discrete method on a published set of DNA mixtures. • We compute likelihood ratios for contributors as well as for non-contributors. • For mixtures with replicates almost the same LR's were found for all contributors. • For mixtures without replicates the information increase for contributors obtained by the continuous method is larger, unless the contributors is one of several minor donors. • False positive rates were very low with both methods. Abstract We present the results of a comparison of likelihood ratios obtained from DNA mixture data, obtained either by a peak height model (EuroForMix) and by a discrete model, using probabilities of dropout and integrating over them (MixKin). We use the mixture data that have been published in [1] and were made publicly available. We show that, for mixtures for which replicate analyses were available in this set, there is almost no difference in weight of evidence, suggesting that the additional information in the peak heights is limited. On the other hand, for mixtures for which only a single replicate is available, the weight of evidence for true donors can be substantially higher with the peak height model, especially so for major donors and also for minor donors provided they are the only minor contributor to the mixture. False positive rates were very low with both methods, but for related non-contributors the risk of a false positive is much higher with the discrete method unless kinship is taken into account in the likelihood ratio calculation. [ABSTRACT FROM AUTHOR]

Highlights • Postmortem swabs transferred to FTA® cards can streamline disaster victim identifications. • Total Body Score (TBS) can be a quick field assessment to evaluate the degree of body decomposition. • No relationship exists between Accumulated Degree Days (ADD) and DNA degradation. • A TBS of 13 and below consistently yielded a full profile regardless of the sampling season or ADD. Abstract In mass fatality events, the need to identify large numbers of deceased persons using DNA can be a significant drain on already overburdened forensic practitioners, both in the field setting and the laboratory. The laboratory may be required to extract DNA from a variety of postmortem sample types, family or direct reference samples related to the missing, and perform matching of these results in a short period of time. While most forensic institutions are well equipped to handle both family and direct reference samples, postmortem samples such as bone or heterogeneous tissue samples can be difficult for labs to analyze. We have devised an easily deployable, efficient, and inexpensive method for collecting postmortem DNA samples on commercially available DNA preservation cards (“FTA®” cards). FTA® cards are already widely used in forensic labs and are convenient for shipping due to their small volume and stability at room temperature. We evaluated the suitability of a protocol involving swabbing of incisions made on cadavers and sample deposition onto FTA® cards over various postmortem intervals and under different environmental conditions. Each trial took place during a different point in the calendar year to evaluate the effects of seasonal weather patterns and temperature on decomposition, DNA yield, and rates of DNA degradation. To further account for the effects of seasonality (temperature and humidity), the progression of body decomposition was recorded following the Total Body Score (TBS) method [1]. DNA degradation was assessed either through STR amplification of 1.2 mm FTA punches or DNA extraction from 3.0 mm punches followed by real-time PCR quantification and STR amplification and genotyping. No consistent relationship was observed between postmortem interval and DNA degradation. Instead, the TBS score, which captures the stage of body decomposition, was shown to correlate well with DNA quantity. A TBS of 15 and below consistently yielded strong partial or full profiles (20 STR loci and Amelogenin using the PowerPlex 21 System) from all individuals from either 1.2 mm or 3.0 mm punches. Transfer of sample swabs to FTA cards is shown to be a simple and effective method for both field and laboratory operations over a range of conditions that can be evaluated by field forensic practitioners based on a body decomposition score. The approach could be beneficially integrated into mass fatality response plans. [ABSTRACT FROM AUTHOR]

Highlights • A new method to determine the shedder status of an individual is presented that is simple to perform. • The use of a DNA staining dye can visualise the presence of cellular material and allow real-time collection of the cellular material to a swab head. • Heavy, intermediate and light shedders were with little variation between marks deposited by an individual’s left and right thumbs. • Correlations can be drawn between the amount of cellular material deposited and the resulting DNA profiles. • By comparing the cellular material in a thumbprint and the resulting DNA profile, an individual’s shedder status can be more accurately determined than previously possible. Abstract All previous examinations of the shedder status of individuals have been based on conclusions inferred from the amount of DNA deposited by donors after they have held an object for a fixed period of time. In all interpretations of shedder status experiments have involved a range uncertainties, especially in regards to results arising from studies carried out in different laboratories. These apply to the efficiency of the swab collecting DNA from the item touched, the amount of DNA left on the swab after attempts to recover it, and the percentage loss of DNA during the lysis and extraction processes. No previous study has attempted to mitigate these uncertainties or verify how much of the DNA deposited was collected through swabbing, how much DNA present on the swab was recovered or how much DNA is lost during the extraction process. We present a study that accurately measures the deposition, collection and amplification of DNA deposited by a range of donors allowing for an accurate determination of the shedder status of individuals. Eleven donors were asked to wash their hands and then deposit a thumbprint onto glass slides by making pressure for 15 seconds 0, 15, 60 and 180 minutes after handwashing. Both left and right thumbs were used and all testing was performed in triplicate. Measurement of the quantity of cellular material deposited on the slides was carried out using DiamondTM Nucleic Acid Dye and fluorescence microscopy on each of 264 thumbprints. Fluorescence microscopy was then used to demonstrate that all the DNA present on the slides was recovered by the swabbing operations and then direct PCR, using the Identifiler™ Plus kit, was used to ensure that none of the DNA present on swabs was lost during DNA profiling. The combination of using a DNA binding dye and direct PCR allowed an accurate means of measuring the extent to which individuals exhibit different extents of shedding. This small study, 11 donors, showed that individuals fell into one of three distinct groups: heavy, intermediate, and light shedders, regardless of the hand used. [ABSTRACT FROM AUTHOR]

The effect of oligomer length on the vibrational mode coupling and energy relaxation mechanisms of AT-rich DNA oligomers in double- and single-stranded conformations has been investigated using two-dimensional infrared spectroscopy. Vibrational coupling of modes of the DNA bases to the symmetric stretching vibration of the backbone phosphate group was observed for oligomers long enough to form duplex-DNA structures. The coupling was lost upon melting of the duplex. No significant effect of oligomer length or DNA secondary structure was found on either the timescale for vibrational relaxation of the base modes or the mechanism, which was consistent with a cascade process from base modes to intermediate modes, some of which are located on the deoxyribose group, and subsequently to the phosphate backbone. The study shows that vibrational coupling between base and backbone requires formation of the double-helix structure while vibrational energy management is an inherent property of the nucleotide. [ABSTRACT FROM AUTHOR]

Radiotherapy has been used for more than a hundred years as a local tumor treatment. The occurrence of systemic antitumor effects manifesting as regression of tumors outside of the irradiated field (abscopal effect) was occasionally observed but deemed too rare and unpredictable to be a therapeutic goal. This has changed with the advent of immunotherapy. Remarkable systemic effects have been observed in patients receiving radiotherapy to control tumors that were progressing during immune checkpoint blockade, stimulating interest in using radiation to overcome primary and acquired cancer resistance to immunotherapy. Here, we review the immunological mechanisms that are responsible for the ability of focal radiation to promote antitumor T cell responses that mediate tumor rejection and, in some cases, result in systemic effects. [ABSTRACT FROM AUTHOR]

Abstract: High‐energy radiation is used in combination with radiosensitizing therapeutics to treat cancer. The most common radiosensitizers are halogenated nucleosides and cisplatin derivatives, and recently also metal nanoparticles have been suggested as potential radiosensitizing agents. The radiosensitizing action of these compounds can at least partly be ascribed to an enhanced reactivity towards secondary low‐energy electrons generated along the radiation track of the high‐energy primary radiation, or to an additional emission of secondary reactive electrons close to the tumor tissue. This is referred to as physico‐chemical radiosensitization. In this Concept article we present current experimental methods used to study fundamental processes of physico‐chemical radiosensitization and discuss the most relevant classes of radiosensitizers. Open questions in the current discussions are identified and future directions outlined, which can lead to optimized treatment protocols or even novel therapeutic concepts. [ABSTRACT FROM AUTHOR]

In this work, we studied the fragmentation dynamics of 2-deoxy-d-ribose (DR) in solution that arises from the double ionization of a water molecule in its primary hydration shell. This process was modeled in the framework of ab initio molecular dynamics. The charge unbalanced in the solvent molecules produces a Coulomb explosion with the consequent release of protons with kinetic energy in the few electronvolts range, which collide with the surrounding molecules in solution inducing further chemical reactions. In particular, we observe proton collisions with the solute molecule DR, which leads to a complete ring opening. In DNA, damage to the DR moiety may lead to DNA strand breaking. This mechanism can be understood as one of the possible steps in the radiation-induced fragmentation of DNA chains. [ABSTRACT FROM AUTHOR]

In genome-wide association studies of complex diseases, many risk polymorphisms are found to lie in non-coding DNA and likely confer risk through allele-dependent differences in gene regulatory elements. However, because distal regulatory elements can alter gene expression at various distances on linear DNA, the identity of relevant genes is unknown for most risk loci. In Parkinson's disease, at least some genetic risk is likely intrinsic to a neuronal subpopulation of cells in the brain regions affected. In order to compare neuron-relevant methods of pairing risk polymorphisms to target genes as well as to further characterize a single-cell model of a neurodegenerative disease, we used the portionally-dopaminergic, neuronal, mesencephalic-derived cell line LUHMES to dissect differentiation-specific mechanisms of gene expression. We compared genome-wide gene expression in undifferentiated and differentiated cells with genome-wide histone H3K27ac and CTCF-bound regions. Whereas promoters and CTCF binding were largely consistent between differentiated and undifferentiated cells, enhancers were mostly unique. We matched the differentiation-specific appearance or disappearance of enhancers with changes in gene expression and identified 22,057 enhancers paired with 6388 differentially expressed genes by proximity. These enhancers are enriched with at least 13 transcription factor response elements, driving a cluster of genes involved in neurogenesis. We show that differentiated LUHMES cells, but not undifferentiated cells, show enrichment for PD-risk SNPs. Candidate genes for these loci are largely unrelated, though a subset is linked to synaptic vesicle cycling and transport, implying that PD-related disruption of these pathways is intrinsic to dopaminergic neurons. [ABSTRACT FROM AUTHOR]

Taxonomic and systematic studies of bamboos are traditionally based on floral morphology, but this can lead to difficulties in identification because of the irregular reproductive cycle of the bamboos. To overcome such problems several molecular-marker approaches have been used. In this study, eight species of the woody bamboos belonging to the genus Dendrocalamus were employed. For each species, DNA samples of 20 individual plants from different localities were isolated and then pooled to make eight bulks of DNA. Fifty RAPD primers were used to screen all bulked DNA samples. Only five primers yielded consistent and reproducible RAPD band patterns across all 160 individuals. The amplicons were present among five species of Dendrocalamus, but were absent in the other three species. They were cloned, sequenced and subsequently, five pairs of SCAR primers were designed. All SCAR primers were combined in multiplex PCR reactions to unequivocally discriminate five species of Dendrocalamus. [ABSTRACT FROM AUTHOR]

Poly(ADP-ribose) polymerase inhibitors (PARPis) are DNA-damaging agents that trap PARP-DNA complexes and interfere with DNA replication. Three PARPis - olaparib, niraparib, and rucaparib - were recently approved by the FDA for the treatment of breast and ovarian cancers. These PARPis, along with 2 others (talazoparib and veliparib), are being evaluated for their potential to treat additional malignancies, including prostate cancers. While lack of PARP-1 confers high resistance to PARPis, it has not been established whether or not the levels of PARP-1 directly correlate with tumor response. In this issue of the JCI, Makvandi and coworkers describe an approach to address this question using [18F]FluorThanatrace, an [18F]-labeled PARP-1 inhibitor, for PET. The tracer was taken up by patient tumor tissue and appeared to differentiate levels of PARP-1 expression; however, future studies should be aimed at determining if this tracer can be used to stratify patient response to PARPi therapy. [ABSTRACT FROM AUTHOR]

DNA damage kills dry-heated spores of Bacillus subtilis, but dry-heattreatment effects on spore germination and outgrowth have not been studied. This is important, since if dry-heat-killed spores germinate and undergo outgrowth, toxic proteins could be synthesized. Here, Raman spectroscopy and differential interference contrast microscopy were used to study germination and outgrowth of individual dry-heat-treated B. subtilis and Bacillus megaterium spores. The major findings in this work were as follows: (i) spores dry-heat-treated at 140°C for 20 min lost nearly all viability but retained their Ca2+-dipicolinic acid (CaDPA) depot; (ii) in most cases, dry-heat treatment increased the average times and variability of all major germination events in B. subtilis spore germination with nutrient germinants or CaDPA, and in one nutrient germination event with B. megaterium spores; (iii) B. subtilis spore germination with dodecylamine, which activates the spore CaDPA release channel, was unaffected by dry-heat treatment; (iv) these results indicate that dry-heat treatment likely damages spore proteins important in nutrient germinant recognition and cortex peptidoglycan hydrolysis, but not CaDPA release itself; and (v) analysis of single spores incubated on nutrient-rich agar showed that while dry-heat-treated spores that are dead can complete germination, they cannot proceed into outgrowth and thus not to vegetative growth. The results of this study provide new information on the effects of dry heat on bacterial spores and indicate that dry-heat sterilization regimens should produce spores that cannot outgrow and thus cannot synthesize potentially dangerous proteins. IMPORTANCE Much research has shown that high-temperature dry heat is a promising means for the inactivation of spores on medical devices and spacecraft decontamination. Dry heat is known to kill Bacillus subtilis spores by DNA damage. However, knowledge about the effects of dry-heat treatment on spore germination and outgrowth is limited, especially at the single spore level. In the current work, Raman spectroscopy and differential interference contrast microscopy were used to analyze CaDPA levels in and kinetics of nutrient- and non-nutrient germination of multiple individual dry-heat-treated B. subtilis and Bacillus megaterium spores that were largely dead. The outgrowth and subsequent cell division of these germinated but dead dryheat- treated spores were also examined. The knowledge obtained in this study will help understand the effects of dry heat on spores both on Earth and in space, and indicates that dry heat can be safely used for sterilization purposes. [ABSTRACT FROM AUTHOR]

Abstract: During cancer radiation therapy high‐energy radiation is used to reduce tumour tissue. The irradiation produces a shower of secondary low‐energy (<20 eV) electrons, which are able to damage DNA very efficiently by dissociative electron attachment. Recently, it was suggested that low‐energy electron‐induced DNA strand breaks strongly depend on the specific DNA sequence with a high sensitivity of G‐rich sequences. Here, we use DNA origami platforms to expose G‐rich telomere sequences to low‐energy (8.8 eV) electrons to determine absolute cross sections for strand breakage and to study the influence of sequence modifications and topology of telomeric DNA on the strand breakage. We find that the telomeric DNA 5′‐(TTA GGG)2 is more sensitive to low‐energy electrons than an intermixed sequence 5′‐(TGT GTG A)2 confirming the unique electronic properties resulting from G‐stacking. With increasing length of the oligonucleotide (i.e., going from 5′‐(GGG ATT)2 to 5′‐(GGG ATT)4), both the variety of topology and the electron‐induced strand break cross sections increase. Addition of K+ ions decreases the strand break cross section for all sequences that are able to fold G‐quadruplexes or G‐intermediates, whereas the strand break cross section for the intermixed sequence remains unchanged. These results indicate that telomeric DNA is rather sensitive towards low‐energy electron‐induced strand breakage suggesting significant telomere shortening that can also occur during cancer radiation therapy. [ABSTRACT FROM AUTHOR]

In this work, we have successfully designed a novel biosensor based on DNA hybridization for ultrasensitive detection of NOS terminator gene sequences (NOSt). This biosensor was synthesized by connecting single-stranded capture DNA (sDNA)-labeled graphene oxide quantum dots (GOQDs) (QDs-sDNA) as fluorescent probe and graphene oxide (GO) as quencher. The detection principle based on hybridization combinations can occur between QDs-sDNA and complementary target DNA; moreover, QDs-sDNA can bind to GO with significantly higher affinity than QDs-dsDNA. In the absence of complementary target DNA, QDs-sDNA was absorbed onto the surface of GO, and the fluorescence of QDs-sDNA was quenched due to fluorescent resonant energy transfer. In the presence of a complementary target DNA, its hybridization with QDs-sDNA formed QDs-dsDNA, which cannot be adsorbed to the GO surface and this leads to reduced quenching. By comparing the fluorescence intensity of QDs-sDNA and QDs-dsDNA in the presence of GO, we can achieve target DNA detection. Thus, rapid, simple, sensitive, efficient, and eco-friendly detection of NOSt was realized. This biosensor had a detection limit of 0.008 nM and a linear range of 0.05–50 nM. Moreover, this sensor can selectivity detect target DNA compared with random and single-base-mismatched sequences, and was successfully applied to the determination target DNA sequences in biological fluids directly. This sensor can be applied to detect other target DNA sequences by simply changing the types of sDNA coupled to the GOQDs. [ABSTRACT FROM AUTHOR]

A recombinant DNA in which the interferon αA (IFN-αA) gene sequence is integrated into a loop region of the gene coding thermostable lichenase was constructed. This approach of insertion fusion with thermostable lichenase is advantageous in terms of increasing the solubility, stability, and production of the fusion partner in soluble form in general and in the periplasm of bacterial cells in particular. Thus, the insertion of IFN-αA into the loop (53 a.a.) of thermostable lichenase from Clostridium thermocellum resulted in effective expression of the soluble form of the recombinant protein in the periplasm of Escherichia coli without any compromise in biological activity of IFN-αA, while the thermostable lichenase retained its ability for functional folding without dramatic loss of its basic activity and thermostability. [ABSTRACT FROM AUTHOR]

Accumulation of mutations in mitochondrial DNA leads to the development of severe, currently untreatable diseases. The contribution of these mutations to aging and progress of neurodegenerative diseases is actively studied. Elucidation of DNA repair mechanisms in mitochondria is necessary for both developing approaches to the therapy of diseases caused by mitochondrial mutations and understanding specific features of mitochondrial genome functioning. Mitochondrial DNA repair systems have become a subject of extensive studies only in the last decade due to development of molecular biology methods. DNA repair systems of mammalian mitochondria appear to be more diverse and effective than it had been thought earlier. Even now, one may speak about the existence of mitochondrial mechanisms for the repair of single-and double-stranded DNA lesions. Homologous recombination also takes place in mammalian mitochondria, although its functional significance and molecular mechanisms remain obscure. In this review, I describe DNA repair systems in mammalian mitochondria, such as base excision repair (BER) and microhomology-mediated end joining (MMEJ) and discuss a possibility of existence of mitochondrial DNA repair mechanisms otherwise typical for the nuclear DNA, e.g., nucleotide excision repair (NER), mismatch repair (MMR), homologous recombination, and classical non-homologous end joining (NHEJ). I also present data on the mechanisms for coordination of the nuclear and mitochondrial DNA repair systems that have been actively studied recently. [ABSTRACT FROM AUTHOR]