DNA Species Research Articles

SARS-CoV-2-induced autophagy dysregulation may cause neuronal dysfunction in COVID-19.

SARS-CoV-2-induced autophagy dysregulation may cause neuronal dysfunction in COVID-19.

DNA barcoding of Iranian leeches (Annelida: Clitellata: Hirudinida)

The diversity of freshwater leeches (Clitellata; Hirudinida) of Iran was estimated by employing both DNA barcoding and species delimitation methods. Phylogenetic relationships of arhynchobdellid (including Hirudinidae, Praobdellidae, Haemopidae, and Erpobdellidae) and rhynchobdellid (Glossiphoniidae) leeches were reconstructed, based on the cytochrome c oxidase subunit I (COI) locus, using both new sequence data and those available from GenBank. Our results suggest that each of Helobdella stagnalis (Linnaeus, 1758), Glossiphonia concolor (Aphathy, 1888), Erpobdella borisi Cichocka & Bielecki, 2015, Dina lineata (O.F. Müller, 1774), Hirudo orientalis Utevsky and Trontelj, 2005, Haemopis sanguisuga (Linnaeus, 1758), Limnatis paluda (Tennent, 1859), and two unidentified species of Dina and Trocheta (these did not find species-level matches in GenBank) are present in Iran. A potential case of phenotypic change in response to ecological adaptation was observed in E. borisi insofar as two genetically identical sub- and super-terranean morphotypes were distinguished. The glossiphoniids of Iran and Europe are admixed in the phylogenetic tree, revealing low COI variation and no divergence within species between the continents for these taxa.

Facile design of multifunction-integrated linear oligonucleotide probe with multiplex amplification effect for label-free and highly sensitive GMO biosensing

Well-defined structures and compositions of nucleic acids afford oligonucleotide probes with unique chemical properties and biological functions for various biosensing applications. Herein, a unique and special oligonucleotide probe, named multifunction-integrated linear oligonucleotide probe (MI-LOP), was facile designed and reported for label-free and turn-on fluorescent detection of the codon component of genetically modified organisms (GMOs). The MI-LOP contains four different functional regions including recognition of target, serving as polymerization template, and creating polymerization primer-linked G-quadruplex (PP-G-quadruplex). Without the aid of any other oligonucleotides, the introduction of target DNA can make each function of the MI-LOP executed one-by-one, during which the species of target DNA, target analogue, and PP-G-quadruplex can be cyclically utilized and in turn induce a multiplex signal amplification responsible for substantial collection of the G-quadruplex moieties under isothermal conditions. The stable G-quadruplexes can combine with N-methyl mesoporphyrin IX (NMM) and function as efficient fluorescence light-up probes, rapidly leading to a dramatic increase in the fluorescence intensity for the amplified detection of the target codon component. Our results strongly demonstrate that the developed MI-LOP with multiplex amplification effect confers the sensing strategy a high sensitivity and specificity for quantitative and qualitative detection of the target codon. And it has also been successfully applied for analyzing target codon in the complex extractions of soybean. The achievements highlight the significance of using oligonucleotide probes as promising analytical tools to promote the basic biochemical research and help in food and environmental analysis.

Significance of Circulating Cell-Free DNA Species in Non-Alcoholic Fatty Liver Disease

The pathogenetic mechanisms involved in the progression of non-alcoholic fatty liver disease (NAFLD) have not been completely elucidated, while the significance of circulating cell-free DNA (cf-DNA) species has been rarely evaluated in NAFLD. Herein, we assessed the serum levels of cf-DNA species in NAFLD patients and investigated their potential associations with patients’ characteristics and severity of liver disease. Forty-nine adult patients with NAFLD of any stage were included in this cohort study. Cf-DNA was isolated from patients’ sera and the levels of several distinct cf-DNA species including total cf-DNA, gene-coding cf-DNA, Alu repeat sequences, mitochondrial DNA copies and 5-methyl-2′-deoxycytidine were determined. Cirrhotic compared to non-cirrhotic patients had significantly lower serum levels of cf-DNA and RNAse P coding DNA as well as higher expression of 5-methyl-2′-deoxycytidine. After adjustment for the significant clinico-epidemiological factors, lower serum levels of cf-DNA or RNAse P were independently associated with the presence of cirrhosis. Serum levels of total and gene-coding DNA are associated with the presence of cirrhosis in NAFLD patients regardless of clinical or epidemiological parameters and may therefore be used as a screening tool for NAFLD progression.

Aquatic Organisms Research with DNA Barcodes

Since their inception, DNA barcodes have become a powerful tool for understanding the biodiversity and biology of aquatic species, with multiple applications in diverse fields such as food security, fisheries, environmental DNA, conservation, and exotic species detection. Nevertheless, most aquatic ecosystems, from marine to freshwater, are understudied, with many species disappearing due to environmental stress, mostly caused by human activities. Here we highlight the progress that has been made in studying aquatic organisms with DNA barcodes, and encourage its further development in assisting sustainable use of aquatic resources and conservation.

Recombination efficiency measurement by real-time PCR: A strategy to evaluate ParA-mediated minicircle production

Minicircles (MCs) are DNA molecules that are produced in Escherichia coli by replicating a parental plasmid (PP) and inducing its site-specific intramolecular recombination into miniplasmid (MP; containing the prokaryotic backbone) and MC molecules (comprised by the eukaryotic cassette). The determination of the recombination efficiency and the monitoring of PP, MC and MP species during processing and in the final product are critical aspects of MC manufacturing. This work describes a real-time PCR method for the specific identification of PP, MP or MC that uses sets of primers specific for each species. The method was evaluated using artificial mixtures of (i) PP and MP, (ii) PP and MC and (iii) MP and MC that were probed for all three DNA molecules. The ratio of molecules of each DNA species in these mixtures were determined with differences lower than 10% relatively to the expected ratio of the species in 90% of the mixtures. Next, the recombination efficiency was successfully estimated by analysing pre-purified DNA samples obtained from cell cultures. A standard deviation < 2% was obtained between replicas and results closely correlated with those obtained by densitometry analysis of agarose gels. Further optimization is required to determine recombination efficiency directly from whole cells.

Evidence of cryptic diversity in freshwater Macrobrachium prawns from Indochinese riverine systems revealed by DNA barcode, species delimitation and phylogenetic approaches.

The diversity of Indochinese prawns in genus Macrobrachium is enormous due to the habitat diversification and broad tributary networks of two river basins: the Chao Phraya and the Mekong. Despite long-standing interest in SE-Asian decapod diversity, the subregional Macrobrachium fauna is still not yet comprehensively clarified in terms of taxonomic identification or genetic diversification. In this study, integrative taxonomic approaches including morphological examination, DNA barcoding, and molecular species delimitation were used to emphasize the broad scale systematics of Macrobrachium prawns in Indochina. Twenty-seven nominal species were successfully re-verified by traditional and molecular taxonomy. Barcode gap analysis supported broad overlapping of species boundaries. Taxonomic ambiguity of several deposited samples in the public database is related to inter- and intraspecific genetic divergence as indicated by BOLD discordance. Diagnostic nucleotide positions were found in six Macrobrachium species. Eighteen additional putative lineages are herein assigned using the consensus of species delimitation methods. Genetic divergence indicates the possible existence of cryptic species in four morphologically complex and wide-ranging species: M. lanchesteri, M. niphanae, M. sintangense, and some members of the M. pilimanus group. The geographical distribution of some species supports the connections and barriers attributed to paleo-historical events of SE-Asian rivers and land masses. Results of this study show explicitly the importance of freshwater ecosystems in Indochinese subregions, especially for the Mekong River Basin due to its high genetic diversity and species composition found throughout its tributaries.

10 STARTUPS TO WATCH 2021: Early-stage companies advance precision medicine by applying AI, clinical genomics, and other new technologies

10 STARTUPS TO WATCH 2021: Early-stage companies advance precision medicine by applying AI, clinical genomics, and other new technologies

The novel Ideal Symmetry Genetic Code table – Common purine-pyrimidine symmetry net for all RNA and DNA species

Ever since Nirenberg’s discovery in 1961 in which codons code individual amino acids, numerous scientists searched for symmetries within the genetic code. The standard genetic code (SGC) table is an alphabetic artificial construct based on the U-C-A-G ordering of nucleotides without natural symmetries. Up to the present, complete symmetry in the genetic code has not been found, leaving doubt as to whether the symmetrical nature as the protector of order even exists. Our novel Ideal Symmetry Genetic Code (ISyGC) table reflects a unique fundamental physicochemical purine-pyrimidine symmetry net for all more than thirty known variations of nuclear and mitochondrial genetic codes. The nuclear genetic code for RNA and DNA viruses also contains the same purine-pyrimidine symmetry net. We show that the ISyGC table leads to automatic transformation into a DNA sequence akin to the 5′3 codon and 3′5 anticodon patterns. As a result of purine-pyrimidine symmetries between codons in the ISyGC table, algorithms of the first two bases as well of the third base of codons show how tRNA cognate anticodons can recognize synonymous codons during mRNA decoding. We show that the ISyGC purine-pyrimidine net with its physicochemical properties represents an evolutionary common “frozen accident” at the onset of each genetic code creation and RNA to DNA evolution. As such, during all of evolution the unique fundamental purine-pyrimidine symmetry net of all genetic codes remains unchangeable. In this way, evolution is a road paved with symmetries.

Plasmonic Ag nanocube enhanced SERS biosensor for sensitive detection of oral cancer DNA based on nicking endonuclease signal amplification and heated electrode

Surface-enhanced Raman scattering (SERS) biosensing has attracted great attention due to its multiplexed capability, high sensitivity, and rapid readout. In this study, plasmonic Ag nanocube (AgNC) enhanced SERS biosensing platform was proposed by coupling nicking endonuclease-assisted target recycling signal amplification with heated electrode for sensitive detection of DNA species related to oral cancer. SERS tags were manufactured by signal DNA (sDNA) conjugating onto the surfaces of AgNCs, and 4-mercaptobenzoic acid (4-MBA) assembled on the surfaces of AgNC/sDNA as a Raman reporter. The sharp feature of edges and corners of AgNCs with high electromagnetic field provided SER hot spots with high quality, and stronger SERS signal could be achieved. In addition, the activity of nicking endonuclease Nt.BstNBI could be greatly enhanced by elevating the temperature of the surface of the heated Au electrode (HAuE) during Nt.BstNBI-assisted target recycling amplification process, resulting in dramatic amplification of target DNA detection. All of these contributed to the high sensitivity of the developed SERS biosensor. A limit of detection (LOD) of 3.1 fM (S/N = 3) could be obtained with an electrode temperature of 45 °C. This proposed SERS biosensor was anticipated to act as a promising biosensing platform for early clinical diagnosis of oral cancer.

Degradation modeling of water environmental DNA: Experiments on multiple DNA sources in pond and seawater

AbstractEnvironmental DNA (eDNA) analysis methods have been developed to detect organism distribution and abundance/biomass in various environments. eDNA degradation is critical for eDNA evaluation. However, the dynamics and mechanisms of eDNA degradation are largely unknown, especially when considering different eDNA sources, for example, cells and fragmental DNA. We experimentally evaluated the degradation rates of eDNA derived from multiple sources, including fragmental DNA (internal PCR control [IPC]), free cells (from Oncorhynchus kisutch), and resident species. We conducted the experiment with pond and seawater to evaluate the differences between freshwater and marine habitats. We quantified the eDNA copies of free cells, fragmental DNA, and resident species (Cyprinus carpio in the pond and Trachurus japonicus in the sea). We found that eDNA derived from both cells and fragmental DNA decreased exponentially in both the sea and pond samples. The degradation of eDNA from resident species showed similar behavior to the cell‐derived eDNA. We evaluated three degradation models with different assumptions and degradation steps and found that a simple exponential model was effective in most cases. Our findings on cell‐ and fragmental DNA‐derived eDNA provide fundamental information about the eDNA degradation process and can be applied to quantify eDNA behavior in natural environments.

Green or white? Morphology, ancient DNA, and the identification of archaeological North American Pacific Coast sturgeon

Sturgeon (Acipenser spp.) have long been an important resource for people living in several parts of the world, including the Northwest Coast of North America. Two sturgeon species occur on the Oregon Coast, white (A. transmontanus) and green (A. medirostris), but morphological similarities between the species have generally prevented osteological identification beyond genus. This lack of species identification hinders evaluation of potential changes in sturgeon range or distribution through time, human food preference or encounter rates, and possible size changes in archaeological sturgeon. We analyzed 1770 sturgeon remains from the Par-Tee site (35CLT20) on the northern Oregon Coast and performed ancient DNA species identification on 30 elements to test morphological distinguishability between species. Ancient DNA analysis indicated that both green and white sturgeon were harvested at the site, but identification to species based on morphology is problematic. Sturgeon were an important fish at several sites in the Northwest Coast and at Par-Tee were a component of a broad-based fishery focused on a range of large and small species. Our study emphasizes the importance of ancient DNA analysis of zooarchaeological assemblages to help address important questions about species composition and abundance.

The complete chloroplast genome of Stauntonia chinensis and compared analysis revealed adaptive evolution of subfamily Lardizabaloideae species in China

BackgroundStauntonia chinensis DC. belongs to subfamily Lardizabaloideae, which is widely grown throughout southern China. It has been used as a traditional herbal medicinal plant, which could synthesize a number of triterpenoid saponins with anticancer and anti-inflammatory activities. However, the wild resources of this species and its relatives were threatened by over-exploitation before the genetic diversity and evolutionary analysis were uncovered. Thus, the complete chloroplast genome sequences of Stauntonia chinensis and comparative analysis of chloroplast genomes of Lardizabaloideae species are necessary and crucial to understand the plastome evolution of this subfamily.ResultsA series of analyses including genome structure, GC content, repeat structure, SSR component, nucleotide diversity and codon usage were performed by comparing chloroplast genomes of Stauntonia chinensis and its relatives. Although the chloroplast genomes of eight Lardizabaloideae plants were evolutionary conserved, the comparative analysis also showed several variation hotspots, which were considered as highly variable regions. Additionally, pairwise Ka/Ks analysis showed that most of the chloroplast genes of Lardizabaloideae species underwent purifying selection, whereas 25 chloroplast protein coding genes were identified with positive selection in this subfamily species by using branch-site model. Bayesian and ML phylogeny on CCG (complete chloroplast genome) and CDs (coding DNA sequences) produced a well-resolved phylogeny of Lardizabaloideae plastid lineages.ConclusionsThis study enhanced the understanding of the evolution of Lardizabaloideae and its relatives. All the obtained genetic resources will facilitate future studies in DNA barcode, species discrimination, the intraspecific and interspecific variability and the phylogenetic relationships of subfamily Lardizabaloideae.

Isotopic Evidence for Long-Distance Connections of the AD Thirteenth-Century Promontory Caves Occupants

The Promontory caves (Utah) and Franktown Cave (Colorado) contain high-fidelity records of short-term occupations by groups with material culture connections to the Subarctic/Northern Plains. This research uses Promontory and Franktown bison dung, hair, hide, and bone collagen to establish local baseline carbon isotopic variability and identify leather from a distant source. The ankle wrap of one Promontory Cave 1 moccasin had a δ13C value that indicates a substantial C4component to the animal's diet, unlike the C3diets inferred from 171 other Promontory and northern Utah bison samples. We draw on a unique combination of multitissue isotopic analysis, carbon isoscapes, ancient DNA (species and sex identification), tissue turnover rates, archaeological contexts, and bison ecology to show that the high δ13C value was not likely a result of local plant consumption, bison mobility, or trade. Instead, the bison hide was likely acquired via long-distance travel to/from an area of abundant C4grasses far to the south or east. Expansive landscape knowledge gained through long-distance associations would have allowed Promontory caves inhabitants to make well-informed decisions about directions and routes of movement for a territorial shift, which seems to have occurred in the late thirteenth century.

Cellular Metabolites Regulate Central Nucleic Acid Sensing Pathways.

Detection of pathogen-derived DNA or RNA species by cellular nucleic acid sensors prompts release of anti-microbial interferons and cytokines. In contrast to their protective anti-microbial functions, inappropriate or excessive activation of nucleic acid sensors can cause inflammatory diseases. Nucleic acid sensing is therefore tightly controlled by regulatory factors acting through both transcriptional and post-transcriptional mechanisms. Recently, it has become clearer that metabolic pathways—previously thought to be unconnected with immune responses—can influence nucleic acid sensing. This regulation can be observed when immune system cells undergo metabolic reprogramming in response to stimulation with pathogen-associated molecular patterns such as lipopolysaccharide from gram negative bacteria. Metabolic reprogramming leads to accumulation and secretion of metabolites, which have been mostly viewed as end-products of processes providing cellular energy and building blocks. However, metabolites have now been identified as important regulators of nucleic acid sensing. This mini-review aims to outline current knowledge on regulation of central nucleic acid sensing pathways by metabolites during metabolic reprogramming.

Implications of Endogenous Retroelements in the Etiopathogenesis of Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease. While its etiology remains elusive, current understanding suggests a multifactorial process with contributions by genetic, immunologic, hormonal, and environmental factors. A hypothesis that combines several of these factors proposes that genomic elements, the L1 retrotransposons, are instrumental in SLE pathogenesis. L1 retroelements are transcriptionally activated in SLE and produce two proteins, ORF1p and ORF2p, which are immunogenic and can drive type I interferon (IFN) production by producing DNA species that activate cytosolic DNA sensors. In addition, these two proteins reside in RNA-rich macromolecular assemblies that also contain well-known SLE autoantigens like Ro60. We surmise that cells expressing L1 will exhibit all the hallmarks of cells infected by a virus, resulting in a cellular and humoral immune response similar to those in chronic viral infections. However, unlike exogenous viruses, L1 retroelements cannot be eliminated from the host genome. Hence, dysregulated L1 will cause a chronic, but perhaps episodic, challenge for the immune system. The clinical and immunological features of SLE can be at least partly explained by this model. Here we review the support for, and the gaps in, this hypothesis of SLE and its potential for new diagnostic, prognostic, and therapeutic options in SLE.

Zebularine elevates STING expression and enhances cGAMP cancer immunotherapy in mice

DNA methylation abnormality is closely related to tumor occurrence and development. Chemical inhibitors targeting DNA methyltransferase (DNMTis) have been used in treating cancer. However, the impact of DNMTis on antitumor immunity has not been well elucidated. In this study, we show that zebularine (a demethylating agent) treatment of cancer cells led to increased levels of interferon response in a cyclic guanosine monophosphate-AMP (cGAMP) synthase (cGAS)- and stimulator of interferon genes (STING)-dependent manner. This treatment also specifically sensitized the cGAS-STING pathway in response to DNA stimulation. Incorporation of zebularine into genomic DNA caused demethylation and elevated expression of a group of genes, including STING. Without causing DNA damage, zebularine led to accumulation of DNA species in the cytoplasm of treated cells. In syngeneic tumor models, administration of zebularine alone reduced tumor burden and extended mice survival. This effect synergized with cGAMP and immune checkpoint blockade therapy. The efficacy of zebularine was abolished in nude mice and in cGAS-/- or STING-/- mice, indicating its dependency on host immunity. Analysis of tumor cells indicates upregulation of interferon-stimulated genes (ISGs) following zebularine administration. Zebularine promoted infiltration of CD8 Tcells and natural killer (NK) cells into tumor and therefore suppressed tumor growth. This study unveils the role of zebularine in sensitizing the cGAS-STING pathway to promote anti-tumor immunity and provides the foundation for further therapeutic development.

On-site genetic analysis for species identification using lab-on-a-chip.

This paper presents a microfluidic device capable of performing genetic analysis on dung samples to identify White Rhinoceros (Ceratotherium simum). The development of a microfluidic device, which can be used in the field, offers a portable and cost‐effective solution for DNA analysis and species identification to aid conservation efforts. Optimization of the DNA extraction processes produced equivalent yields compared to conventional kit‐based methods within just 5 minutes. The use of a color‐changing loop‐mediated isothermal amplification reaction for simultaneous detection of the cytochrome B sequence of C. simum enabled positive results to be obtained within as little as 30 minutes. Field testing was performed at Knowsley Safari to demonstrate real‐world applicability of the microfluidic device for testing of biological samples.

Electrical DNA Sequence Mapping Using Oligodeoxynucleotide Labels and Nanopores.

Identifying DNA species is crucial for diagnostics. For DNA identification, single-molecule DNA sequence mapping is an alternative to DNA sequencing toward fast point-of-care testing, which traditionally relies on targeting and labeling DNA sequences with fluorescent labels and readout using optical imaging methods. A nanopore is a promising sensor as a complement to optical mapping with advantages of electric measurement suitable for portable devices and potential for high resolution. Here, we demonstrate a high-resolution nanopore-based DNA sequence mapping by labeling specific short sequence motifs with oligodeoxynucleotides (ODNs) using DNA methyltransferase (MTase) and detecting them using nanopores. We successfully detected ODNs down to the size of 11 nucleotides without introducing extra reporters and resolved neighboring sites with a distance of 141 bp (∼48 nm) on a single DNA molecule. To accurately locate the sequence motif positions on DNA, a nanopore data analysis method is proposed by considering DNA velocity change through nanopores and using ensemble statistics to translate the time-dependent signals to the location information. Our platform enables high-resolution detection of small labels on DNA and high-accuracy localization of them for DNA species identification in an all-electrical format. The method presents an alternative to optical techniques relying on fluorescent labels and is promising for miniature-scale integration for diagnostic applications.

Triplex and other DNA motifs show motif-specific associations with mitochondrial DNA deletions and species lifespan

The “theory of resistant biomolecules” posits that long-lived species show resistance to molecular damage at the level of their biomolecules. Here, we test this hypothesis in the context of mitochondrial DNA (mtDNA) as it implies that predicted mutagenic DNA motifs should be inversely correlated with species maximum lifespan (MLS).First, we confirmed that guanine-quadruplex and direct repeat (DR) motifs are mutagenic, as they associate with mtDNA deletions in the human major arc of mtDNA, while also adding mirror repeat (MR) and intramolecular triplex motifs to a growing list of potentially mutagenic features. What is more, triplex motifs showed disease-specific associations with deletions and an apparent interaction with guanine-quadruplex motifs.Surprisingly, even though DR, MR and guanine-quadruplex motifs were associated with mtDNA deletions, their correlation with MLS was explained by the biased base composition of mtDNA. Only triplex motifs negatively correlated with MLS even after adjusting for body mass, phylogeny, mtDNA base composition and effective number of codons.Taken together, our work highlights the importance of base composition for the comparative biogerontology of mtDNA and suggests that future research on mitochondrial triplex motifs is warranted.