A novel method of microsatellite genotyping-by-sequencing using individual combinatorial barcoding

This study examines the potential of next-generation sequencing based ‘genotyping-by-sequencing’ (GBS) of microsatellite loci for rapid and cost-effective genotyping in large-scale population genetic studies. The recovery of individual genotypes from large sequence pools was achieved by PCR-incorporated combinatorial barcoding using universal primers. Three experimental conditions were employed to explore the possibility of using this approach with existing and novel multiplex marker panels and weighted amplicon mixture. The GBS approach was validated against microsatellite data generated by capillary electrophoresis. GBS allows access to the underlying nucleotide sequences that can reveal homoplasy, even in large datasets and facilitates cross laboratory transfer. GBS of microsatellites, using individual combinatorial barcoding, is potentially faster and cheaper than current microsatellite approaches and offers better and more data.

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Next Gen Pop Gen: implementing a high-throughput approach to population genetics in boarfish ( Capros aper )

Royal Society Open Science ◽

10.1098/rsos.160651 ◽

2016 ◽

Vol 3 (12) ◽

pp. 160651 ◽

Cited By ~ 13

Author(s):

Edward D. Farrell ◽

Jeanette E. L. Carlsson ◽

Jens Carlsson

Keyword(s):

De Novo ◽

Genotyping By Sequencing ◽

Cost Effective ◽

Species Range ◽

Genetic Population ◽

Genetic Studies ◽

Scale Population ◽

Management And Conservation ◽

Generation Sequencing ◽

Insular Populations

The recently developed approach for microsatellite genotyping by sequencing (GBS) using individual combinatorial barcoding was further improved and used to assess the genetic population structure of boarfish ( Capros aper ) across the species' range. Microsatellite loci were developed de novo and genotyped by next-generation sequencing. Genetic analyses of the samples indicated that boarfish can be subdivided into at least seven biological units (populations) across the species' range. Furthermore, the recent apparent increase in abundance in the northeast Atlantic is better explained by demographic changes within this area than by influx from southern or insular populations. This study clearly shows that the microsatellite GBS approach is a generic, cost-effective, rapid and powerful method suitable for full-scale population genetic studies—a crucial element for assessment, sustainable management and conservation of valuable biological resources.

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A Novel Method for the Preparation of Fibrous CeO2–ZrO2–Y2O3 Compacts for Thermochemical Cycles

Crystals ◽

10.3390/cryst11080885 ◽

2021 ◽

Vol 11 (8) ◽

pp. 885

Author(s):

Nicole Knoblauch ◽

Peter Mechnich

Keyword(s):

Large Scale ◽

Preparation Method ◽

Cost Effective ◽

High Porosity ◽

Thermochemical Cycles ◽

Redox Cycles ◽

Redox Kinetics ◽

Direct Infiltration ◽

Novel Method ◽

Functional Components

Zirconium-Yttrium-co-doped ceria (Ce0.85Zr0.13Y0.02O1.99) compacts consisting of fibers with diameters in the range of 8–10 µm have been successfully prepared by direct infiltration of commercial YSZ fibers with a cerium oxide matrix and subsequent sintering. The resulting chemically homogeneous fiber-compacts are sinter-resistant up to 1923 K and retain a high porosity of around 58 vol% and a permeability of 1.6–3.3 × 10−10 m² at a pressure gradient of 100–500 kPa. The fiber-compacts show a high potential for the application in thermochemical redox cycling due its fast redox kinetics. The first evaluation of redox kinetics shows that the relaxation time of oxidation is five times faster than that of dense samples of the same composition. The improved gas exchange due to the high porosity also allows higher reduction rates, which enable higher hydrogen yields in thermochemical water-splitting redox cycles. The presented cost-effective fiber-compact preparation method is considered very promising for manufacturing large-scale functional components for solar-thermal high-temperature reactors.

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Validation of variants using cost effective highresolution melting (HRM) analysis predicted from target re-sequencing in Eucalyptus

Acta Botanica Croatica ◽

10.37427/botcro-2020-019 ◽

2020 ◽

Vol 79 (2) ◽

pp. 105-113

Author(s):

Abdul Bari Muneera Parveen ◽

Divya Lakshmanan ◽

Modhumita Ghosh Dasgupta

Keyword(s):

Next Generation Sequencing ◽

Large Scale ◽

Sequence Data ◽

Cost Effective ◽

Nucleotide Polymorphisms ◽

Next Generation ◽

Time Saving ◽

Hrm Analysis ◽

The Cost ◽

Generation Sequencing

The advent of next-generation sequencing has facilitated large-scale discovery and mapping of genomic variants for high-throughput genotyping. Several research groups working in tree species are presently employing next generation sequencing (NGS) platforms for marker discovery, since it is a cost effective and time saving strategy. However, most trees lack a chromosome level genome map and validation of variants for downstream application becomes obligatory. The cost associated with identifying potential variants from the enormous amount of sequence data is a major limitation. In the present study, high resolution melting (HRM) analysis was optimized for rapid validation of single nucleotide polymorphisms (SNPs), insertions or deletions (InDels) and simple sequence repeats (SSRs) predicted from exome sequencing of parents and hybrids of Eucalyptus tereticornis Sm. ? Eucalyptus grandis Hill ex Maiden generated from controlled hybridization. The cost per data point was less than 0.5 USD, providing great flexibility in terms of cost and sensitivity, when compared to other validation methods. The sensitivity of this technology in variant detection can be extended to other applications including Bar-HRM for species authentication and TILLING for detection of mutants.

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Next-Generation Sequencing Technologies in Blood Group Typing

Transfusion Medicine and Hemotherapy ◽

10.1159/000504765 ◽

2019 ◽

Vol 47 (1) ◽

pp. 4-13 ◽

Cited By ~ 1

Author(s):

Daniel Fürst ◽

Chrysanthi Tsamadou ◽

Christine Neuchel ◽

Hubert Schrezenmeier ◽

Joannis Mytilineos ◽

...

Keyword(s):

Next Generation Sequencing ◽

Blood Group ◽

Large Scale ◽

Cost Effective ◽

Molecular Testing ◽

Blood Group Antigens ◽

Next Generation ◽

Sequencing Technologies ◽

Blood Group Typing ◽

Generation Sequencing

Sequencing of the human genome has led to the definition of the genes for most of the relevant blood group systems, and the polymorphisms responsible for most of the clinically relevant blood group antigens are characterized. Molecular blood group typing is used in situations where erythrocytes are not available or where serological testing was inconclusive or not possible due to the lack of antisera. Also, molecular testing may be more cost-effective in certain situations. Molecular typing approaches are mostly based on either PCR with specific primers, DNA hybridization, or DNA sequencing. Particularly the transition of sequencing techniques from Sanger-based sequencing to next-generation sequencing (NGS) technologies has led to exciting new possibilities in blood group genotyping. We describe briefly the currently available NGS platforms and their specifications, depict the genetic background of blood group polymorphisms, and discuss applications for NGS approaches in immunohematology. As an example, we delineate a protocol for large-scale donor blood group screening established and in use at our institution. Furthermore, we discuss technical challenges and limitations as well as the prospect for future developments, including long-read sequencing technologies.

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STAMP: a multiplex sequencing method for simultaneous evaluation of mitochondrial DNA heteroplasmies and content

10.1101/2020.07.05.188607 ◽

2020 ◽

Author(s):

Xiaoxian Guo ◽

Yiqin Wang ◽

Ruoyu Zhang ◽

Zhenglong Gu

Keyword(s):

Large Scale ◽

Nuclear Dna ◽

Large Population ◽

High Sensitivity ◽

Cost Effective ◽

Cost Effective Method ◽

Sequencing Method ◽

Scale Population ◽

Mtdna Variants ◽

Multiplex Sequencing

ABSTRACTHuman mitochondrial genome (mtDNA) variations, such as mtDNA heteroplasmies (the co-existence of mutated and wild-type mtDNA), have received increasing attention in recent years for their clinical relevance to numerous diseases. But large-scale population studies of mtDNA heteroplasmies have been lagging due to the lack of a labor- and cost-effective method. Here, we present a novel human mtDNA sequencing method called STAMP (sequencing by targeted amplification of multiplex probes) for measuring mtDNA heteroplasmies and content in a streamlined workflow. We show that STAMP has high mapping rates to mtDNA, deep coverage of unique reads, and high tolerance to sequencing and PCR errors when applied to human samples. STAMP also has high sensitivity and low false positive rates in identifying artificial mtDNA variants at fractions as low as 0.5% in genomic DNA samples. We further extend STAMP, by including nuclear DNA-targeting probes, to enable assessment of relative mtDNA content in the same assay. The high cost-effectiveness of STAMP, along with the flexibility of using it for measuring various aspects of mtDNA variations, will accelerate the research of mtDNA heteroplasmies and content in large population cohorts, and in the context of human diseases and aging.

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STAMP: a multiplex sequencing method for simultaneous evaluation of mitochondrial DNA heteroplasmies and content

NAR Genomics and Bioinformatics ◽

10.1093/nargab/lqaa065 ◽

2020 ◽

Vol 2 (4) ◽

Author(s):

Xiaoxian Guo ◽

Yiqin Wang ◽

Ruoyu Zhang ◽

Zhenglong Gu

Keyword(s):

Large Scale ◽

Nuclear Dna ◽

Large Population ◽

High Sensitivity ◽

Cost Effective ◽

Cost Effective Method ◽

Sequencing Method ◽

Scale Population ◽

Mtdna Variants ◽

Multiplex Sequencing

Abstract Human mitochondrial genome (mtDNA) variations, such as mtDNA heteroplasmies (the co-existence of mutated and wild-type mtDNA), have received increasing attention in recent years for their clinical relevance to numerous diseases. But large-scale population studies of mtDNA heteroplasmies have been lagging due to the lack of a labor- and cost-effective method. Here, we present a novel human mtDNA sequencing method called STAMP (sequencing by targeted amplification of multiplex probes) for measuring mtDNA heteroplasmies and content in a streamlined workflow. We show that STAMP has high-mapping rates to mtDNA, deep coverage of unique reads and high tolerance to sequencing and polymerase chain reaction errors when applied to human samples. STAMP also has high sensitivity and low false positive rates in identifying artificial mtDNA variants at fractions as low as 0.5% in genomic DNA samples. We further extend STAMP, by including nuclear DNA-targeting probes, to enable assessment of relative mtDNA content in the same assay. The high cost-effectiveness of STAMP, along with the flexibility of using it for measuring various aspects of mtDNA variations, will accelerate the research of mtDNA heteroplasmies and content in large population cohorts, and in the context of human diseases and aging.

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Sequence-Selective Recognition of Extended-Spectrum β-Lactamase GES-2 by a Competitive, Peptide Nucleic Acid-Based Multiplex PCR Assay

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.9.3402-3406.2004 ◽

2004 ◽

Vol 48 (9) ◽

pp. 3402-3406 ◽

Cited By ~ 6

Author(s):

Gerhard F. Weldhagen

Keyword(s):

Nucleic Acid ◽

Multiplex Pcr ◽

Peptide Nucleic Acid ◽

Large Scale ◽

Cost Effective ◽

Coding Region ◽

Screening Programs ◽

Extended Spectrum ◽

Multiplex Pcr Assay ◽

Novel Method

ABSTRACT Extended-spectrum β-lactamases (ESBLs) in Pseudomonas aeruginosa, such as GES-2, which compromises the efficacy of imipenem, tend to be geographically restricted. The CC-to-AA base pair substitution at positions 493 and 494 of the bla GES-2-coding region distinguishes this ESBL from bla GES-1 and the bla IBC-type genes, making it an ideal target for the development of a novel sequence-specific, peptide nucleic acid (PNA)-based multiplex PCR detection method. By using two primer pairs in conjunction with a PNA probe, this method provided an accurate means of identification of bla GES-2 compared to standard PCR and gene sequencing techniques when it was used to test 100 P. aeruginosa clinical isolates as well as previously published, well-described control strains encompassing all presently known genes in the bla GES-IBC ESBL family. This novel method has the potential to be used in large-scale, cost-effective screening programs for specific or geographically restricted ESBLs.

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Genotyping-by-Sequencing (GBS): A Novel, Efficient and Cost-Effective Genotyping Method for Cattle Using Next-Generation Sequencing

PLoS ONE ◽

10.1371/journal.pone.0062137 ◽

2013 ◽

Vol 8 (5) ◽

pp. e62137 ◽

Cited By ~ 104

Author(s):

Marcos De Donato ◽

Sunday O. Peters ◽

Sharon E. Mitchell ◽

Tanveer Hussain ◽

Ikhide G. Imumorin

Keyword(s):

Next Generation Sequencing ◽

Genotyping By Sequencing ◽

Cost Effective ◽

Next Generation ◽

Genotyping Method ◽

Generation Sequencing

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TOGGLe, a flexible framework for easily building complex workflows and performing robust large-scale NGS analyses

10.1101/245480 ◽

2018 ◽

Cited By ~ 4

Author(s):

Christine Tranchant-Dubreuil ◽

Sébastien Ravel ◽

Cécile Monat ◽

Gautier Sarah ◽

Abdoulaye Diallo ◽

...

Keyword(s):

Population Structure ◽

High Performance ◽

Large Scale ◽

Genotyping By Sequencing ◽

Whole Genome ◽

Third Generation ◽

Third Generation Sequencing ◽

Sequencing Method ◽

Flexible Framework ◽

Generation Sequencing

ABSTRACTThe advent of NGS has intensified the need for robust pipelines to perform high-performance automated analyses. The required softwares depend on the sequencing method used to produce raw data (e.g. Whole genome sequencing, Genotyping By Sequencing, RNASeq) as well as the kind of analyses to carry on (GWAS, population structure, differential expression). These tools have to be generic and scalable, and should meet the biologists needs.Here, we present the new version of TOGGLe (Toolbox for Generic NGS Analyses), a simple and highly flexible framework to easily and quickly generate pipelines for large-scale second- and third-generation sequencing analyses, including multi-sample and multi-threading support. TOGGLe is a workflow manager designed to be as effortless as possible to use for biologists, so the focus can remain on the analyses. Pipelines are easily customizable and supported analyses are reproducible and shareable. TOGGLe is designed as a generic, adaptable and fast evolutive solution, and has been tested and used in large-scale projects on various organisms. It is freely available at http://toggle.southgreen.fr/, under the GNU GPLv3/CeCill-C licenses) and can be deployed onto HPC clusters as well as on local machines.

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Ancestry informative alleles captured with reduced representation library sequencing in Theobroma cacao

10.1101/407387 ◽

2018 ◽

Author(s):

Jaime A. Osorio-Guarín ◽

Corey R. Quackenbush ◽

Omar E. Cornejo

Keyword(s):

Large Scale ◽

Demographic History ◽

Genotyping By Sequencing ◽

Cost Effective ◽

Snp Discovery ◽

Nucleotide Polymorphism ◽

Single Nucleotide ◽

Breeding Programs ◽

Reduced Representation ◽

Reduced Representation Library

AbstractAs the source of chocolate, cacao has become one of the most important crops in the world. The identification of molecular markers to understand the demographic history, genetic diversity and population structure plays a pivotal role in cacao breeding programs. Here, we report the use of a modified genotyping-by-sequencing (GBS) approach for large-scale single nucleotide polymorphism (SNP) discovery and allele ancestry mapping. We identified 12,357 bi-allelic SNPs after filtering, of which, 7,009 variants were ancestry informative. The GBS approach proved to be rapid, cost-effective, and highly informative for ancestry assignment in this species.

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