scholarly journals A Fast and Accessible Method for the Isolation of RNA, DNA, and Protein to Facilitate the Detection of SARS-CoV-2

2020 ◽  
Author(s):  
Jose Carlos Ponce-Rojas ◽  
Michael S. Costello ◽  
Duncan A. Proctor ◽  
Kenneth S. Kosik ◽  
Maxwell Z. Wilson ◽  
...  
Keyword(s):  
Low Cost ◽  
Rna Extraction ◽  
Genetic Material ◽  
Simple Method ◽  
Dna And Rna ◽  
Alternative Method ◽  
Accessible Method ◽  
Comparable Performance ◽  
Viral Envelopes ◽  
Common Laboratory

AbstractManagement of the COVID-19 pandemic requires widespread SARS-CoV-2 testing. A main limitation for widespread SARS-CoV-2 testing is the global shortage of essential supplies, among these, RNA extraction kits. The need for commercial RNA extraction kits places a bottleneck on tests that detect SARS-CoV-2 genetic material, including PCR-based reference tests. Here we propose an alternative method we call PEARL (Precipitation Enhanced Analyte RetrievaL) that addresses this limitation. PEARL uses a lysis solution that disrupts cell membranes and viral envelopes while simultaneously providing conditions suitable for alcohol-based precipitation of RNA, DNA, and proteins. PEARL is a fast, low-cost, and simple method that uses common laboratory reagents and offers comparable performance to commercial RNA extraction kits. PEARL offers an alternative method to isolate host and pathogen nucleic acids and proteins to streamline the detection of DNA and RNA viruses, including SARS-CoV-2.

scholarly journals A Fast and Accessible Method for the Isolation of RNA, DNA, and Protein to Facilitate the Detection of SARS-CoV-2

2020 ◽  
Author(s):  
Jose Carlos Ponce-Rojas ◽  
Michael S. Costello ◽  
Duncan A. Proctor ◽  
Kenneth S. Kosik ◽  
Maxwell Z. Wilson ◽  
...  
Keyword(s):  
Low Cost ◽  
Rna Extraction ◽  
Genetic Material ◽  
Simple Method ◽  
Dna And Rna ◽  
Alternative Method ◽  
Accessible Method ◽  
Comparable Performance ◽  
Viral Envelopes ◽  
Common Laboratory

Management of the COVID-19 pandemic requires widespread SARS-CoV-2 testing. A main limitation for widespread SARS-CoV-2 testing is the global shortage of essential supplies, among these, RNA extraction kits. The need for commercial RNA extraction kits places a bottleneck on tests that detect SARS-CoV-2 genetic material, including PCR-based reference tests. Here we propose an alternative method we call PEARL (Precipitation Enhanced Analyte RetrievaL) that addresses this limitation. PEARL uses a lysis solution that disrupts cell membranes and viral envelopes while simultaneously providing conditions suitable for alcohol-based precipitation of RNA, DNA, and proteins. PEARL is a fast, low-cost, and simple method that uses common laboratory reagents and offers comparable performance to commercial RNA extraction kits. PEARL offers an alternative method to isolate host and pathogen nucleic acids and proteins to streamline the detection of DNA and RNA viruses, including SARS-CoV-2.

Killing two birds with one stone: simultaneous extraction of DNA and RNA from activated sludge biomass

1999 ◽  
Vol 45 (3) ◽  
pp. 269-272 ◽  
Author(s):  
Zhongtang Yu ◽  
William W Mohn
Keyword(s):  
Activated Sludge ◽  
Ammonium Acetate ◽  
Rna Extraction ◽  
Cell Lysis ◽  
Biomass Density ◽  
Simple Method ◽  
Dna And Rna ◽  
Acid Recovery ◽  
Bacterial Aggregate ◽  
Soil And Sediment

DNA and RNA are usually extracted from activated sludge samples using two separate methods developed for soil and sediment samples. However, activated sludge differs from soil and sediment in at least three aspects: high biomass density, low humic acid content, and the presence of bacterial aggregate flocs. Taking these characteristics into consideration, we developed a simple and rapid method allowing simultaneous DNA and RNA extraction from activated sludge samples. This method combines (i) mini-bead beating, which is most efficient in breaking bacterial aggregate flocs and cells, (ii) protection of RNA with diethyl pyrocarbonate, and (iii) precipitation of impurities with ammonium acetate. Phenol/chloroform extraction and column purification are not necessary. The resulting DNA and RNA are suitable for PCR and reverse transcriptase - PCR, respectively. The efficiencies of cell lysis and nucleic acid recovery were high enough to permit detection by PCR of 102cells/mL of mixed liquor. By simultaneously extracting both DNA and RNA from a single sample, this method eliminates variability in cell lysis between extraction of DNA and RNA using two different methods. This extraction method is rapid, and within 1 h, one person can process four or more samples. This simple method makes it easier to analyze a large number of activated sludge samples.Key words: activated sludge, DNA, extraction, PCR, RNA.

scholarly journals Sample collection/stabilization and DNA/RNA extraction from swab samples for microbiome or metagenome analyses

BioTechniques ◽  
2021 ◽  
Author(s):  
David Navarro ◽  
A González ◽  
A Saiz O y Odriozola
Keyword(s):  
Microbial Communities ◽  
Rna Extraction ◽  
Genetic Material ◽  
New Product ◽  
Storage Medium ◽  
Sample Collection ◽  
Bacterial Composition ◽  
Good Preservation ◽  
Dna And Rna ◽  
And Storage

Good preservation and storage are essential to preserving microorganisms’ genetic material in microbial communities from wide array of sample inputs and accurately represent the bacterial composition for further analysis and applications. The objective is to develop a proper preservation and storage medium to preserve DNA and RNA from those microorganisms. DANAGEN-BIOTED has developed a new product to deal with this problem. Click on the To read the full Application forum, click on the View Article button above and download the PDF.

A simple method to construct a low-cost immunosensor based on a dithiol-functionalized polydopamine platform

2021 ◽  
Author(s):  
Bo Liu ◽  
Luanying Yang ◽  
Gang Wang ◽  
Sha He ◽  
Xiaobo Wang ◽  
...  
Keyword(s):  
Detection Limit ◽  
Linear Response ◽  
Low Cost ◽  
Covalent Immobilization ◽  
The Self ◽  
Simple Method ◽  
Low Detection Limit

A simple and low-cost electrochemical CEA immunosensor was investigated via the self-polymerization of dopamine and a dithiol compound spacer for the covalent immobilization of antibodies. The designed CEA immunosensor exhibited a linear response and a low detection limit.

scholarly journals A set of simple methods for detection and extraction of laminarinase

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ananthamurthy Koteshwara ◽  
Nancy V. Philip ◽  
Jesil Mathew Aranjani ◽  
Raghu Chandrashekhar Hariharapura ◽  
Subrahmanyam Volety Mallikarjuna
Keyword(s):  
Ammonium Sulfate ◽  
Ammonium Sulphate ◽  
Gold Standard ◽  
Direct Detection ◽  
Low Cost ◽  
Direct Analysis ◽  
Reducing Sugar ◽  
Ammonium Sulfate Precipitation ◽  
Simple Method ◽  
Functional Step

AbstractA carefully designed ammonium sulfate precipitation will simplify extraction of proteins and is considered to be a gold standard among various precipitation methods. Therefore, optimization of ammonium sulfate precipitation can be an important functional step in protein purification. The presence of high amounts of ammonium sulphate precludes direct detection of many enzymatically active proteins including reducing sugar assays (e.g. Nelson-Somogyi, Reissig and 3,5-dinitrosalicylic acid methods) for assessing carbohydrases (e.g. laminarinase (β (1–3)-glucanohydrolase), cellulases and chitinases). In this study, a simple method was developed using laminarin infused agarose plate for the direct analysis of the ammonium sulphate precipitates from Streptomyces rimosus AFM-1. The developed method is simple and convenient that can give accurate results even in presence of ammonium sulfate in the crude precipitates. Laminarin is a translucent substrate requiring the use of a stain to visualize the zones of hydrolysis in a plate assay. A very low-cost and locally available fluorescent optical fabric brightener Tinopal CBS-X has been used as a stain to detect the zones of hydrolysis. We also report simple methods to prepare colloidal chitin and cell free supernatant in this manuscript.

A Very Low-Cost, Labor-Efficient, and Simple Method to Block Scattered Ultraviolet Light in PDMS Microfluidic Devices by Inserting Aluminum Foil Strips

2018 ◽  
Vol 11 (1) ◽  
Author(s):  
Shuo Wang ◽  
Peter Shankles ◽  
Scott Retterer ◽  
Yong Tae Kang ◽  
Chang Kyoung Choi
Keyword(s):  
Soft Lithography ◽  
Uv Light ◽  
Low Cost ◽  
Microfluidic Devices ◽  
Aluminum Foil ◽  
Material Synthesis ◽  
Simple Method ◽  
Micro Particles ◽  
Complex Shapes ◽  
The Cost

Abstract Opto-microfluidic methods have advantages for manufacturing complex shapes or structures of micro particles/hydrogels. Most of these microfluidic devices are made of polydimethylsiloxane (PDMS) by soft lithography because of its flexibility of designing and manufacturing. However, PDMS scatters ultraviolet (UV) light, which polymerizes the photocrosslinkable materials at undesirable locations and clogs the microfluidic devices. A fluorescent dye has previously been employed to absorb the scattered UV light and shift its wavelength to effectively solve this issue. However, this method is limited due to the cost of the materials (tens of dollars per microchip), the time consumed on synthesizing the fluorescent material and verifying its quality (two to three days). More importantly, significant expertise on material synthesis and characterization is required for users of the opto-microfluidic technique. The cost of preliminary testing on multiple iterations of different microfluidic chip designs would also be excessive. Alternatively, with a delicate microchannel design, we simply inserted aluminum foil strips (AFS) inside the PDMS device to block the scattered UV light. By using this method, the UV light was limited to the exposure region so that the opto-microfluidic device could consistently generate microgels longer than 6 h. This is a nearly cost- and labor-free method to solve this issue.

scholarly journals A simple interferometric method to measure the calibration factor and displacement amplification in piezoelectric flextensional actuators

2010 ◽  
Vol 21 (6) ◽  
pp. 577-587 ◽  
Author(s):  
Francisco de Assis Andrade Barbosa ◽  
Gilder Nader ◽  
Ricardo Tokio Higuti ◽  
Cláudio Kitano ◽  
Emílio Carlos Nelli Silva
Keyword(s):  
Modulation Depth ◽  
Low Cost ◽  
Michelson Interferometer ◽  
Laser Interferometry ◽  
Optimization Method ◽  
Calibration Factor ◽  
Simple Method ◽  
Interferometric Method ◽  
Established Technique

Laser interferometry is a well-established technique for the characterization of piezoelectric actuators. In this work, by using a low cost Michelson interferometer, the measurement of the calibration factor and the displacement amplification of a novel piezoelectric flextensional actuator (PFA), designed by using the topology optimization method, is performed. A simple method, based on small phase modulation depth when the PFA is driven by a triangular waveform, allows the absolute interferometer calibration. The free-displacement of the PFA for various drive voltages is measured and the displacement amplification is determined. The linearity and frequencyresponse of the PFA are evaluated up to 20 kHz

scholarly journals Guidance for laboratories performing molecular pathology for cancer patients

2014 ◽  
Vol 67 (11) ◽  
pp. 923-931 ◽  
Author(s):  
Ian A Cree ◽  
Zandra Deans ◽  
Marjolijn J L Ligtenberg ◽  
Nicola Normanno ◽  
Anders Edsjö ◽  
...  
Keyword(s):  
Molecular Pathology ◽  
Rna Extraction ◽  
Internal Audit ◽  
Turnaround Time ◽  
Careful Consideration ◽  
Internal Quality ◽  
Molecular Testing ◽  
Pathology Report ◽  
Sufficient Information ◽  
Dna And Rna

Molecular testing is becoming an important part of the diagnosis of any patient with cancer. The challenge to laboratories is to meet this need, using reliable methods and processes to ensure that patients receive a timely and accurate report on which their treatment will be based. The aim of this paper is to provide minimum requirements for the management of molecular pathology laboratories. This general guidance should be augmented by the specific guidance available for different tumour types and tests. Preanalytical considerations are important, and careful consideration of the way in which specimens are obtained and reach the laboratory is necessary. Sample receipt and handling follow standard operating procedures, but some alterations may be necessary if molecular testing is to be performed, for instance to control tissue fixation. DNA and RNA extraction can be standardised and should be checked for quality and quantity of output on a regular basis. The choice of analytical method(s) depends on clinical requirements, desired turnaround time, and expertise available. Internal quality control, regular internal audit of the whole testing process, laboratory accreditation, and continual participation in external quality assessment schemes are prerequisites for delivery of a reliable service. A molecular pathology report should accurately convey the information the clinician needs to treat the patient with sufficient information to allow for correct interpretation of the result. Molecular pathology is developing rapidly, and further detailed evidence-based recommendations are required for many of the topics covered here.

scholarly journals Tuning Dielectric Loss of SiO2@CNTs for Electromagnetic Wave Absorption

Nanomaterials ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 2636
Author(s):  
Fenghui Cao ◽  
Jia Xu ◽  
Xinci Zhang ◽  
Bei Li ◽  
Xiao Zhang ◽  
...  
Keyword(s):  
Electromagnetic Wave ◽  
High Performance ◽  
Low Cost ◽  
Impedance Matching ◽  
3D Structure ◽  
Carbon Matrix ◽  
Current Approach ◽  
Matrix Composites ◽  
Simple Method ◽  
Absorption Performance

We developed a simple method to fabricate SiO2-sphere-supported N-doped CNTs (NCNTs) for electromagnetic wave (EMW) absorption. EMW absorption was tuned by adsorption of the organic agent on the precursor of the catalysts. The experimental results show that the conductivity loss and polarization loss of the sample are improved. Meanwhile, the impedance matching characteristics can also be adjusted. When the matching thickness was only 1.5 mm, the optimal 3D structure shows excellent EMW absorption performance, which is better than most magnetic carbon matrix composites. Our current approach opens up an effective way to develop low-cost, high-performance EMW absorbers.

scholarly journals Glyoxal fixation facilitates transcriptome analysis after antigen staining and cell sorting by flow cytometry

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0240769
Author(s):  
Prasanna Channathodiyil ◽  
Jonathan Houseley
Keyword(s):  
Flow Cytometry ◽  
Transcriptome Analysis ◽  
Mammalian Cells ◽  
Single Cells ◽  
Rna Extraction ◽  
Cyclin B1 ◽  
Immunofluorescent Staining ◽  
Rna Seq ◽  
Simple Method ◽  
Staining Methods

A simple method for extraction of high quality RNA from cells that have been fixed, stained and sorted by flow cytometry would allow routine transcriptome analysis of highly purified cell populations and single cells. However, formaldehyde fixation impairs RNA extraction and inhibits RNA amplification. Here we show that good quality RNA can be readily extracted from stained and sorted mammalian cells if formaldehyde is replaced by glyoxal—a well-characterised fixative that is widely compatible with immunofluorescent staining methods. Although both formaldehyde and glyoxal efficiently form protein-protein crosslinks, glyoxal does not crosslink RNA to proteins nor form stable RNA adducts, ensuring that RNA remains accessible and amenable to enzymatic manipulation after glyoxal fixation. We find that RNA integrity is maintained through glyoxal fixation, permeabilisation with methanol or saponin, indirect immunofluorescent staining and flow sorting. RNA can then be extracted by standard methods and processed into RNA-seq libraries using commercial kits; mRNA abundances measured by poly(A)+ RNA-seq correlate well between freshly harvested cells and fixed, stained and sorted cells. We validate the applicability of this approach to flow cytometry by staining MCF-7 cells for the intracellular G2/M-specific antigen cyclin B1 (CCNB1), and show strong enrichment for G2/M-phase cells based on transcriptomic data. Switching to glyoxal fixation with RNA-compatible staining methods requires only minor adjustments of most existing staining and sorting protocols, and should facilitate routine transcriptomic analysis of sorted cells.

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