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143 results for “DNA extraction”
Figure 1 in Application of the NucliSENS easyMAG system for nucleic acid extraction: optimization of DNA extraction for molecular diagnosis of parasitic and fungal diseases
Figure 1. Study design.
Fig.1 in Protocol Optimization For Genomic Dna Extraction And Rapd-Pcr Of Alien Ponto-Caspian Amphipod Pontogammarus Robustoides
Fig.1. Localities of sampling sities in the Latvian reservoirs.
An unsupervised deep learning framework with variational autoencoders for genome-wide DNA methylation analysis and biologic feature extraction applied to breast cancer
<p>Supplemental data for the paper titled "An unsupervised deep learning framework with variational autoencoders for genome-wide DNA methylation analysis and biologic feature extraction applied to breast cancer"</p>
FIGURE 2 in DNA extracted from museum specimens of the 19 century provides a taxonomic resolution on the identity of the characid fish Psalidodon jequitinhonhae (Ostariophysi: Characiformes)
FIGURE 2 | Lectotype of Tetragonopterus rutilus jequitinhonhae, NMW 57760:2, 67.52 mm SL.
Optimizing DNA extraction protocols for the diet analysis of a baleen whale (Eubalaena australis)
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Data for: Environmental DNA storage and extraction method affects detectability for multiple aquatic invasive species
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Data from: Comparative analysis of DNA extraction methods to study the body surface microbiota of insects: a case study with ant cuticular bacteria
High-throughput sequencing of the 16S rRNA gene has considerably helped revealing the essential role of bacteria living on insect cuticles in the ecophysiology and behavior of their hosts. However, our understanding of host-cuticular microbiota feedbacks remains hampered by the difficulties to working with low bacterial DNA quantities as in individual insect cuticle samples, which are more prone to molecular biases and contaminations. Herein, we conducted a methodological benchmark on the cuticular bacterial loads retrieved from two Neotropical ant species of different body size and ecology: Atta cephalotes (~15 mm) and Pseudomyrmex penetrator (~5 mm). We evaluated the richness and composition of the cuticular microbiota, as well as the amount of biases and contamination produced by four DNA extraction protocols. We also addressed how bacterial communities' characteristics would be affected by the number of individuals or individual body size used for DNA extraction. Most extraction methods yielded similar results in term of bacterial diversity and composition for A. cephalotes (~15 mm). In contrast, greater amounts of artifactual sequences and contaminations, as well as noticeable differences in bacterial communities' characteristics were observed between the extraction methods for P. penetrator (~5 mm). We also found that large (~15 mm) and small (~5 mm) A. cephalotes individuals harbor different bacterial communities. Our benchmark hence suggests that cuticular microbiota of single insect individuals can be reliably retrieved provided that blank controls, appropriate data cleaning, and standardization of individual body size are considered in the experiment.
Data from: High-throughput sequencing of nematode communities from total soil DNA extractions
Background: Nematodes are extremely diverse and numbers of species are predicted to be more than a million. Studies on nematode diversity are difficult and laborious using standard methods such as identification based on morphology and therefore high-throughput sequencing is an attractive alternative. Generally, primers that have been used for generating amplicons for sequencing are not nematode specific and also amplify other groups such as fungi and plantae. Thus a nematode enrichment step must be included that may introduce biases. Results: An amplification strategy, including a new primer, which selectively amplifies nematodes and other metazoans was developed. When this strategy was tested on DNA templates from a set of 22 agricultural soils, we obtained 64.4 % sequences of nematode origin in total, whereas the remaining sequences were almost entirely metazoan. The nematode sequences were derived from a broad taxonomic range and most sequences were from nematode taxa that have previously been found to be abundant in soil such as Tylenchida, Rhabditida, Dorylaimida, Triplonchida and Araeolaimida. Conclusions: This amplification and sequencing strategy for assessing nematode diversity was demonstrated to be able to collect a broad taxonomy of nematodes without prior enrichment and thus the method will be highly valuable in ecological studies of nematodes. Keywords: nematode, community, next-generation sequencing, SSU, diversity, 18S, rDNA
Data from: Extracting DNA from 'jaws': high yield and quality from archived tiger shark (Galeocerdo cuvier) skeletal material
Archived specimens are highly valuable sources of DNA for retrospective genetic/genomic analysis. However, often limited effort has been made to evaluate and optimize extraction methods, which may be crucial for downstream applications. Here, we assessed and optimized the usefulness of abundant archived skeletal material from sharks as a source of DNA for temporal genomic studies. Six different methods for DNA extraction, encompassing two different commercial kits and three different protocols, were applied to material, so-called bio-swarf, from contemporary and archived jaws and vertebrae of tiger sharks (Galeocerdo cuvier). Protocols were compared for DNA yield and quality using a qPCR approach. For jaw swarf, all methods provided relatively high DNA yield and quality, while large differences in yield between protocols were observed for vertebrae. Similar results were obtained from samples of white shark (Carcharodon carcharias). Application of the optimized methods to 38 museum and private angler trophy specimens dating back to 1912 yielded sufficient DNA for downstream genomic analysis for 68% of the samples. No clear relationships between age of samples, DNA quality and quantity were observed, likely reflecting different preparation and storage methods for the trophies. Trial sequencing of DNA capture genomic libraries using 20 000 baits revealed that a significant proportion of captured sequences were derived from tiger sharks. This study demonstrates that archived shark jaws and vertebrae are potential high-yield sources of DNA for genomic-scale analysis. It also highlights that even for similar tissue types, a careful evaluation of extraction protocols can vastly improve DNA yield.
Evaluation of different methods to extract DNA from serum seeded with methicillin-resistant Staphylococcus aureus
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Raw data of sequencing results of our study: Bovine milk microbiota: Evaluation of different DNA extraction protocols in challenging samples
<p>Clean reads of the repeated milk samples with used Primer Pairs V1V2 and V3V4</p> <p>Raw data of sequencing results (amplicon single variants)</p>
Optimising recovery of DNA from minimally-invasive sampling methods: efficacy of buccal swabs, preservation strategy and DNA extraction approaches for amphibian studies_Dataset_Rscript
<p>Datasets and Rscript associated with paper draft titled: "<span>Optimising recovery of DNA from minimally-invasive sampling methods: efficacy of buccal swabs, preservation strategy and DNA extraction approaches for amphibian studies".</span></p> <p> </p> <p>Abstract: <span>Studies in evolution, ecology and conservation are increasingly based on genetic and genomic inferences. With increased focus on molecular approaches, ethical concerns about destructive or more invasive techniques need to be considered, with a push for minimally invasive sampling to be optimised. Buccal swabs have been increasingly used to collect DNA in a number of taxa, including amphibians.<span> </span>However, DNA yield and purity from swabs is often low, limiting its use. In this study we compare different types of swabs, preservation method and storage, and DNA extraction technique in three case studies to assess the optimal approach for recovering DNA in anurans. Out of the five different types of swab that we tested, Isohelix MS-02 and Rapidry swabs generated higher DNA yields than other swabs. When comparing storage buffers, ethanol is a better preservative than a non-alcoholic alternative. Dried samples resulted in similar or better final DNA yields than ethanol-fixed samples if kept cool. DNA extraction via a Qiagen</span><span>™</span><span> DNeasy Blood and Tissue Kit and McHale’s salting out extraction method resulted in similar DNA yields but the Qiagen</span><span>™</span><span> kit extracts contained less contamination. We also found that samples produce better DNA recovery if frozen as soon as possible after collection. We provide recommendations for sample collection and extraction under different conditions, including budgetary considerations, size of individual sampled, access to cold storage facilities, and DNA extraction methodology. Maximising efficacy of all of these factors for better DNA recovery will allow buccal swabs to be used for genetic and genomic studies in a range of vertebrates.</span></p>
Data from: Choice of capture and extraction methods affect detection of freshwater biodiversity from environmental DNA
Environmental DNA (eDNA) is used to detect biodiversity by the capture, extraction, and identification of DNA shed to the environment. However, eDNA capture and extraction protocols vary widely across studies. This use of different protocols potentially biases detection results and could significantly hinder a reliable use of eDNA to detect biodiversity. We tested whether choice of eDNA capture and extraction protocols significantly influenced biodiversity detection in aquatic systems. We sampled lake and river water, captured and extracted eDNA using six combinations of different protocols with replication, and tested for the detection of four macroinvertebrate species. Additionally, using the same lake water technical replicates, we compared the effect of capture and extraction protocols on metabarcode detections of biodiversity using 16S for eubacteria and cytochrome c oxidase I (COI) for eukaryotes. Protocol combinations for capture and extraction of eDNA significantly influenced DNA yield and number of sequences obtained from next generation sequencing. We found significantly different detection rates of species ranging from zero percent to thirty-three percent. Differences in which protocol combinations produced the highest metabarcoded biodiversity were detected and demonstrate that different protocols are required for different biodiversity targets. Our results highlight that the choice of molecular protocols used for capture and extraction of eDNA from water can strongly affect biodiversity detection. Consideration of biases caused by choice of protocols should lead to a more consistent and reliable molecular workflow for repeatable and increased detection of biodiversity in aquatic communities.
Data from: HyRAD-X, a versatile method combining exome capture and RAD sequencing to extract genomic information from ancient DNA
Over the last decade, protocols aimed at reproducibly sequencing reduced-genome subsets in non-model organisms have been widely developed. Their use is however limited to DNA of relatively high molecular weight. During the last year, several methods exploiting hybridization capture using probes based on RAD-sequencing loci have circumvented this limitation and opened avenues to the study of samples characterized by degraded DNA, such as historical specimens. Here, we present a major update to those methods, namely Hybridization capture from RAD-derived probes obtained from a reduced eXome template (hyRAD-X), a technique applying RAD-sequencing to messenger RNA from one or few fresh specimens to elaborate bench-top produced probes, i.e., a reduced representation of the exome, further used to capture homologous DNA from a samples set. In contrast to previous hybridization-capture methods, the reference catalog on which reads are aligned does not rely on de novo assembly of anonymous RAD-sequencing loci, but on an assembled transcriptome obtained from RNAseq data, thus increasing the accuracy of loci definition and Single-Nucleotide-Polmorphisms (SNP) call, and targeting, specifically, expressed genes. Finally, the capture step of hyRAD-X relies on RNA probes, increasing stringency of hybridization, making it well suited for low-content DNA samples. As a proof of concept, we applied hyRAD-X to subfossil needles from the coniferous tree Abies alba, collected in lake sediments (Origlio, Switzerland) and dating back from 7200-5800 years before present (BP). More specifically we investigated genetic variation before, during, and after an anthropogenic perturbation that caused an abrupt decrease in Abies alba population size, 6500-6200 years BP. HyRAD-X produced a matrix encompassing 524 exome-derived SNPs. Despite a lower observed heterozygosity was observed during the 6.500-6.200 years BP time slice, genetic composition was nearly identical before and after the perturbation, indicating that re-expansion of the population after the decline was driven by autochthonous specimens. To the best of our knowledge, this is the first time a population genomic study incorporating ancient DNA samples of tree subfossils is conducted at a moderate cost using reproducible exome-reduced complexity.
Figure 2 in Successful identification of the final instar nymph of Quintilia carinata (Thunberg) (Hemiptera: Cicadidae) by DNA extraction from the exuvium
Figure 2. Legs of the final instar nymph of Quintilia carinata: (A) Lateral view of left foreleg. acf, accessory tooth of femur; apt, apical tooth of tibia; bt, blade of tibia; f, femur; fc, femoral comb; itf, intermediate tooth of femur; pbt, point of blade of tibia; ptf, posterior tooth of femur; t, trochanter; ti, tibia; ta, tarsus. (B) Distal spines of mesotibia. (C) Distal spines of metatibia. Scale bars: (A) 2 mm, (B) 1 mm, (C) 1 mm.
Figure 1 in Successful identification of the final instar nymph of Quintilia carinata (Thunberg) (Hemiptera: Cicadidae) by DNA extraction from the exuvium
Figure 1. Exuvium of the final instar nymph of Quintilia carinata. (A) Lateral view. (B) Dorsal view of head. Scale bars: (A) 8 mm, (B) 2 mm.
Unlocking the genomes of formalin-fixed freshwater fish specimens: An assessment of factors influencing DNA extraction quantity and quality
<p>DNA quality and quantity metrics (Microsoft Excel worksheet with 3 sheets, one per species).</p> <p>We compared two protocols developed to extract DNA from formalin-fixed tissues using specimens of three freshwater fishes: Southern Brook Lamprey <em>Ichthyomyzon gagei</em>, Slimy Sculpin <em>Cottus cognatus</em>, and Brown Trout <em>Salmo trutta</em>. Extractions were attempted using hot alkali digestion with and without buffer wash pretreatments to compare the DNA concentration, purity, and fragment length of DNA recovered between extraction protocols, tissue types (muscle and caudal fin tissue for Brown Trout and Slimy Sculpin), and preservation periods (5 or 7 years for Southern Brook Lamprey).</p> <p>Four metrics were collected: DNA quantity (measured using Qubit 3.0 instrument with dsDNA high sensitivity kit); DNA purity measured by A260/A280 ratio (measured using NanoDrop 2000 spectrophotometer); DNA purity measured by A260/A230 ratio (measured using NanoDrop 2000 spectrophotometer); and percent fragments >= 300 base pairs (measured using ImageJ). For spectrophotometric absorbance ratios, an A260/A280 ratio of 1.8 and an A260/ A230 ratio of 2.0 were considered to represent “pure” DNA (i.e., free of contaminants). For analyses of DNA purity, deviations from 1.8 for the A260/A280 ratio and from 2.0 for the A260/A230 ratio were used as the response variables, such that larger deviation values corresponded to less-pure DNA.</p> <p>Please see attached README.md file for additional information.</p>
Data from: High-throughput sequencing of nematode communities from total soil DNA extractions
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Data from: Extracellular DNA extraction is a fast, cheap and reliable alternative for multi-taxa surveys based on soil DNA
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Data from: A cost-efficient and simple protocol to enrich prey DNA from extractions of predatory arthropods for large-scale gut content analysis by Illumina sequencing
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