scholarly journals A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample

2017 ◽  
Author(s):  
Lisa Olohan ◽  
Laura-Jayne Gardiner ◽  
Anita Lucaci ◽  
Burkhard Steuernagel ◽  
Brande Wulff ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Cost Effective ◽  
Capture Probe ◽  
Sequence Capture ◽  
Reduction Techniques ◽  
Capture Assay ◽  
Starting Point ◽  
Agilent Sureselect ◽  
Type Data ◽  
Single Capture

AbstractBackgroundBread wheat has a large complex genome that makes whole genome resequencing costly. Therefore, genome complexity reduction techniques such as sequence capture make re-sequencing cost effective. With a high-quality draft wheat genome now available it is possible to design capture probe sets and to use them to accurately genotype and anchor SNPs to the genome. Furthermore, in addition to genetic variation, epigenetic variation provides a source of natural variation contributing to changes in gene expression and phenotype that can be profiled at the base pair level using sequence capture coupled with bisulphite treatment. Here, we present a new 12 Mbp wheat capture probe set, that allows both the profiling of genotype and methylation from the same DNA sample. Furthermore, we present a method, based on Agilent SureSelect Methyl-Seq, that will use a single capture assay as a starting point to allow both DNA sequencing and methyl-seq.ResultsOur method uses a single capture assay that is sequentially split and used for both DNA sequencing and methyl-seq. The resultant genotype and epi-type data is highly comparable in terms of coverage and SNP/methylation site identification to that generated from separate captures for DNA sequencing and methyl-seq. Furthermore, by defining SNP frequencies in a diverse landrace from the Watkins collection we highlight the importance of having genotype data to prevent false positive methylation calls. Finally, we present the design of a new 12 Mbp wheat capture and demonstrate its successful application to re-sequence wheat.ConclusionWe present a cost-effective method for performing both DNA sequencing and methyl-seq from a single capture reaction thus reducing reagent costs, sample preparation time and DNA requirements for these complementary analyses.

scholarly journals Next-generation sequencing in the clinical genetic screening of patients with pheochromocytoma and paraganglioma

2013 ◽  
Vol 2 (2) ◽  
pp. 104-111 ◽  
Author(s):  
Joakim Crona ◽  
Alberto Delgado Verdugo ◽  
Dan Granberg ◽  
Staffan Welin ◽  
Peter Stålberg ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Genetic Screening ◽  
Bioinformatics Analysis ◽  
Cost Effective ◽  
Susceptibility Genes ◽  
Next Generation ◽  
Clinical Genetic ◽  
Cost Effective Method ◽  
Agilent Sureselect ◽  
Generation Sequencing

BackgroundRecent findings have shown that up to 60% of pheochromocytomas (PCCs) and paragangliomas (PGLs) are caused by germline or somatic mutations in one of the 11 hitherto known susceptibility genes: SDHA, SDHB, SDHC, SDHD, SDHAF2, VHL, HIF2A (EPAS1), RET, NF1, TMEM127 and MAX. This list of genes is constantly growing and the 11 genes together consist of 144 exons. A genetic screening test is extensively time consuming and expensive. Hence, we introduce next-generation sequencing (NGS) as a time-efficient and cost-effective alternative.MethodsTumour lesions from three patients with apparently sporadic PCC were subjected to whole exome sequencing utilizing Agilent Sureselect target enrichment system and Illumina Hi seq platform. Bioinformatics analysis was performed in-house using commercially available software. Variants in PCC and PGL susceptibility genes were identified.ResultsWe have identified 16 unique genetic variants in PCC susceptibility loci in three different PCC, spending less than a 30-min hands-on, in-house time. Two patients had one unique variant each that was classified as probably and possibly pathogenic: NF1 Arg304Ter and RET Tyr791Phe. The RET variant was verified by Sanger sequencing.ConclusionsNGS can serve as a fast and cost-effective method in the clinical genetic screening of PCC. The bioinformatics analysis may be performed without expert skills. We identified process optimization, characterization of unknown variants and determination of additive effects of multiple variants as key issues to be addressed by future studies.

scholarly journals Towards Biohydrogen Separation Using Poly(Ionic Liquid)/Ionic Liquid Composite Membranes

Membranes ◽  
2018 ◽  
Vol 8 (4) ◽  
pp. 124 ◽  
Author(s):  
Andreia S.L. Gouveia ◽  
Lucas Ventaja ◽  
Liliana C. Tomé ◽  
Isabel M. Marrucho
Keyword(s):  
Ionic Liquid ◽  
High Performance ◽  
Cost Effective ◽  
Clean Energy ◽  
Composite Membranes ◽  
Feed Pressure ◽  
Starting Point ◽  
Poly Ionic Liquid ◽  
Separation Performances ◽  
H2 Selectivity

Considering the high potential of hydrogen (H2) as a clean energy carrier, the implementation of high performance and cost-effective biohydrogen (bioH2) purification techniques is of vital importance, particularly in fuel cell applications. As membrane technology is a potentially energy-saving solution to obtain high-quality biohydrogen, the most promising poly(ionic liquid) (PIL)–ionic liquid (IL) composite membranes that had previously been studied by our group for CO2/N2 separation, containing pyrrolidinium-based PILs with fluorinated or cyano-functionalized anions, were chosen as the starting point to explore the potential of PIL–IL membranes for CO2/H2 separation. The CO2 and H2 permeation properties at the typical conditions of biohydrogen production (T = 308 K and 100 kPa of feed pressure) were measured and discussed. PIL–IL composites prepared with the [C(CN)3]− anion showed higher CO2/H2 selectivity than those containing the [NTf2]− anion. All the membranes revealed CO2/H2 separation performances above the upper bound for this specific separation, highlighting the composite incorporating 60 wt% of [C2mim][C(CN)3] IL.

An International Campaign for Agricultural and Livestock Genomics (CALG)

2002 ◽  
Vol 06 (24) ◽  
pp. 958-965
Author(s):  
Jun Yu ◽  
Jian Wang ◽  
Huanming Yang
Keyword(s):  
Large Scale ◽  
Cost Effective ◽  
Model Organisms ◽  
Environmental Biology ◽  
Cdna Sequences ◽  
Governmental Agencies ◽  
Technology Innovations ◽  
A Genome ◽  
Starting Point ◽  
The Cost

A coordinated international effort to sequence agricultural and livestock genomes has come to its time. While human genome and genomes of many model organisms (related to human health and basic biological interests) have been sequenced or plugged in the sequencing pipelines, agronomically important crop and livestock genomes have not been given high enough priority. Although we are facing many challenges in policy-making, grant funding, regional task emphasis, research community consensus and technology innovations, many initiatives are being announced and formulated based on the cost-effective and large-scale sequencing procedure, known as whole genome shotgun (WGS) sequencing that produces draft sequences covering a genome from 95 percent to 99 percent. Identified genes from such draft sequences, coupled with other resources, such as molecular markers, large-insert clones and cDNA sequences, provide ample information and tools to further our knowledge in agricultural and environmental biology in the genome era that just comes to its accelerated period. If the campaign succeeds, molecular biologists, geneticists and field biologists from all countries, rich or poor, would be brought to the same starting point and expect another astronomical increase of basic genomic information, ready to convert effectively into knowledge that will ultimately change our lives and environment into a greater and better future. We call upon national and international governmental agencies and organizations as well as research foundations to support this unprecedented movement.

scholarly journals Towards Biohydrogen Separation Using Poly(ionic liquid)/ionic Liquid Composite Membranes

2018 ◽  
Author(s):  
Andreia S.L. Gouveia ◽  
Lucas Ventaja ◽  
Liliana C. Tome ◽  
Isabel M. Marrucho
Keyword(s):  
Ionic Liquid ◽  
High Performance ◽  
Cost Effective ◽  
Clean Energy ◽  
Composite Membranes ◽  
Feed Pressure ◽  
Starting Point ◽  
Poly Ionic Liquid ◽  
Separation Performances ◽  
Permeation Properties

Considering the high potential of hydrogen (H2) as a clean energy carrier, the implementation of high performance and cost-effective biohydrogen (bioH2) purification techniques is of vital importance, particularly in fuel cell applications. In this context, membrane technology is a potentially energy-saving solution to obtain high-quality biohydrogen. The most promising poly(ionic liquid) (PIL) - ionic liquid (IL) composite membranes previously studied by our group for CO2/N2 separation, containing pyrrolidinium-based PILs with fluorinated or cyano-functionalized anions, were chosen as starting point to explore the potential of PIL–IL membranes for CO2/H2 separation. The CO2 and H2 permeation properties at the typical conditions of biohydrogen production (T =308 K and 100 kPa of feed pressure) were measured and discussed. PIL–IL composites prepared with [C(CN)3]– anion showed higher CO2/H2 selectivities and H2 diffusivities compared to those containing [NTf2]– anion. All the membranes revealed CO2/H2 separation performances above the upper bound for this specific separation, highlighting the composite incorporating 60 wt% of [C2mim][C(CN)3] IL.

scholarly journals Deep sequencing of DNA from urine of kidney allograft recipients to estimate donor/recipient-specific DNA fractions

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0249930
Author(s):  
Aziz Belkadi ◽  
Gaurav Thareja ◽  
Darshana Dadhania ◽  
John R. Lee ◽  
Thangamani Muthukumar ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Bacterial Infections ◽  
Human Kidney ◽  
Cost Effective ◽  
Maximum Error ◽  
Standard Test ◽  
Kidney Allograft ◽  
Cost Effective Method ◽  
In Silico Simulation ◽  
End Stage

Kidney transplantation is the treatment of choice for patients with end-stage kidney failure, but transplanted allograft could be affected by viral and bacterial infections and by immune rejection. The standard test for the diagnosis of acute pathologies in kidney transplants is kidney biopsy. However, noninvasive tests would be desirable. Various methods using different techniques have been developed by the transplantation community. But these methods require improvements. We present here a cost-effective method for kidney rejection diagnosis that estimates donor/recipient-specific DNA fraction in recipient urine by sequencing urinary cell DNA. We hypothesized that in the no-pathology stage, the largest tissue types present in recipient urine are donor kidney cells, and in case of rejection, a larger number of recipient immune cells would be observed. Extensive in-silico simulation was used to tune the sequencing parameters: number of variants and depth of coverage. Sequencing of DNA mixture from 2 healthy individuals showed the method is highly predictive (maximum error < 0.04). We then demonstrated the insignificant impact of familial relationship and ethnicity using an in-house and public database. Lastly, we performed deep DNA sequencing of urinary cell pellets from 32 biopsy-matched samples representing two pathology groups: acute rejection (AR, 11 samples) and acute tubular injury (ATI, 12 samples) and 9 samples with no pathology. We found a significant association between the donor/recipient-specific DNA fraction in the two pathology groups compared to no pathology (P = 0.0064 for AR and P = 0.026 for ATI). We conclude that deep DNA sequencing of urinary cells from kidney allograft recipients offers a noninvasive means of diagnosing acute pathologies in the human kidney allograft.

Beyond Textbooks

2019 ◽  
pp. 182-200 ◽  
Author(s):  
Lesley S. J. Farmer
Keyword(s):  
Teaching And Learning ◽  
Online Courses ◽  
Cost Effective ◽  
Open Educational Resources ◽  
Educational Resources ◽  
Academic Needs ◽  
Online Instructors ◽  
Starting Point ◽  
Good Starting Point ◽  
Reference Tool

Textbooks can serve as a good starting point for learning concepts or serve as a reinforcing reference tool for students. However, to address the various academic needs of students, as well as to affirm the richness and depth of the knowledge, skills, and dispositions in online courses, online instructors should complement and supplement textbooks with other resources in various formats. Education has a growing need for digital resources, starting with digital textbooks, and expanding to other kinds of educational resources. Open educational resources, in particular, provide cost-effective and flexible tools for teaching and learning.

scholarly journals Highly multiplexed, fast and accurate nanopore sequencing for verification of synthetic DNA constructs and sequence libraries

2019 ◽  
Vol 4 (1) ◽  
Author(s):  
Andrew Currin ◽  
Neil Swainston ◽  
Mark S Dunstan ◽  
Adrian J Jervis ◽  
Paul Mulherin ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Dna Sequencing ◽  
Cost Effective ◽  
Polymorphism Analysis ◽  
Sequencing Data ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Synthetic Dna ◽  
Design Build ◽  
Hardware Costs

Abstract Synthetic biology utilizes the Design–Build–Test–Learn pipeline for the engineering of biological systems. Typically, this requires the construction of specifically designed, large and complex DNA assemblies. The availability of cheap DNA synthesis and automation enables high-throughput assembly approaches, which generates a heavy demand for DNA sequencing to verify correctly assembled constructs. Next-generation sequencing is ideally positioned to perform this task, however with expensive hardware costs and bespoke data analysis requirements few laboratories utilize this technology in-house. Here a workflow for highly multiplexed sequencing is presented, capable of fast and accurate sequence verification of DNA assemblies using nanopore technology. A novel sample barcoding system using polymerase chain reaction is introduced, and sequencing data are analyzed through a bespoke analysis algorithm. Crucially, this algorithm overcomes the problem of high-error rate nanopore data (which typically prevents identification of single nucleotide variants) through statistical analysis of strand bias, permitting accurate sequence analysis with single-base resolution. As an example, 576 constructs (6 × 96 well plates) were processed in a single workflow in 72 h (from Escherichia coli colonies to analyzed data). Given our procedure’s low hardware costs and highly multiplexed capability, this provides cost-effective access to powerful DNA sequencing for any laboratory, with applications beyond synthetic biology including directed evolution, single nucleotide polymorphism analysis and gene synthesis.

An enzyme-linked immunosorbent assay for the detection of Rathayibacter toxicus, the bacterium involved in annual ryegrass toxicity, in hay

2006 ◽  
Vol 57 (7) ◽  
pp. 731 ◽  
Author(s):  
A. M. Masters ◽  
A. R. Gregory ◽  
R. J. Evans ◽  
J. E. Speijers ◽  
S. S. Sutherland
Keyword(s):  
Monoclonal Antibody ◽  
Immunosorbent Assay ◽  
Specific Antigen ◽  
Cost Effective ◽  
Enzyme Linked Immunosorbent Assay ◽  
Annual Ryegrass ◽  
Capture Assay ◽  
Large Numbers ◽  
Soluble Polysaccharide ◽  
Annual Ryegrass Toxicity

An enzyme-linked immunosorbent assay (ELISA) for Rathayibacter toxicus is described. The development of a monoclonal antibody for a specific antigen from R. toxicus and a polyclonal antibody raised against the same R. toxicus preparation enabled a capture assay format. The assay is specific for a soluble polysaccharide produced by the bacterium and was found to be sensitive enough to detect antigen equivalent to less than one gall per kilogram of hay. The applicability of the assay to samples of pasture or hay is demonstrated. Cost-effective testing of large numbers of samples for the presence of R. toxicus is possible with the ELISA. This will assist stockowners, hay producers, and hay exporters in the management of the risk of annual ryegrass toxicity.

Noise Reduction Retrofit on Historic Structures

1996 ◽  
Vol 1559 (1) ◽  
pp. 26-28
Author(s):  
Suzanne DiGeronimo ◽  
Lewis S. Goodfriend
Keyword(s):  
Noise Reduction ◽  
Cost Effective ◽  
Modern World ◽  
Historic Buildings ◽  
Reduction Techniques ◽  
Sound Levels ◽  
Historic Structures ◽  
Air Intake ◽  
Continuous Use ◽  
Speech Interference

Cost-effective noise reduction techniques for historic buildings can be accomplished affordably and without altering the structure's historic status. Techniques include adding sashes to existing historic windows, replacing windows with visually compatible acoustic units, and using jamb extensions to accommodate manufacturers' standard window units. Wood-truss roofs can be insulated from noise on the interior through the addition of a light-gauge metal framing support with alternating layers of gypsum board, plywood, and batt insulation. Ventilation solutions include a unique over-the-sill air intake system that functions as an acoustic baffle and visually spares a historic facade from random louver punch-throughs. Acoustically rated doors and vestibule construction address noise reduction at entry ways. At an example installation, sound levels were reduced by 12 dB, and speech interference levels were reduced by 18–23 dB. Continuous use is the best method for preservation of historic buildings. Noise reduction, sensibly applied, gives a historic structure occupational viability in the modern world.

Clinical Needs as a Starting Point for Different Strategies in Computational Drug Development

Drug Research ◽  
2018 ◽  
Vol 69 (08) ◽  
pp. 458-466 ◽  
Author(s):  
César Portela
Keyword(s):  
Drug Development ◽  
Translational Research ◽  
Computational Chemistry ◽  
Therapeutic Potential ◽  
Cost Effective ◽  
New Drugs ◽  
Medical Practitioners ◽  
Pathophysiological Mechanisms ◽  
Starting Point ◽  
The Way

AbstractTraditionally, the first step in the development of drugs is the definition of the target, by choice of a biological structure involved in a disease or by recognition of a molecule with some degree of a biological activity that presents itself as druggable and endowed with therapeutic potential. The complexity of the pathophysiological mechanisms of disease and of the structures of the molecules involved creates several challenges in this drug discovery process. These difficulties also come from independent operation of the different parts involved in drug development, with little interaction between clinical practitioners, academic institutions and large pharmaceutical companies. Research in this area is purpose specific, performed by specialized researchers in each field, without major inputs from clinical practitioners on the relevance of such strategy for future therapies. Translational research can shift the way these relationships operate towards a process in which new therapies can be generated by linking experimental discoveries directly to unmet clinical needs. Computational chemistry methods provide valuable insights on experimental findings and pharmacological and pathophysiological mechanisms, allow the virtual construction of new possibilities for the synthesis of new molecular entities, and pave the way for informed cost-effective decisions on expensive research projects. This text focus on the current computational methods used in drug design, how they can be used in a translational research model that starts from clinical practice and research-based theorization by medical practitioners and moves to applied research in a computational chemistry setting, aiming the development of new drugs for clinical use.

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