scholarly journals Deciphering the genetic diversity of landraces with high-throughput SNP genotyping of DNA bulks: methodology and application to the maize 50k array

2020 ◽  
Author(s):  
Mariangela Arca ◽  
Tristan Mary-Huard ◽  
Brigitte Gouesnard ◽  
Aurélie Bérard ◽  
Cyril Bauland ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Throughput ◽  
Large Scale ◽  
Allelic Frequency ◽  
Laboratory Equipment ◽  
Fluorescent Intensity ◽  
Allelic Frequencies ◽  
Wide Range ◽  
Young Leaves ◽  
Scale Characterization

ABSTRACTGenebanks harbor original landraces carrying many original favorable alleles for mitigating biotic and abiotic stresses. Their genetic diversity remains however poorly characterized due to their large within genetic diversity. We developed a high-throughput, cheap and labor saving DNA bulk approach based on SNP Illumina Infinium HD array to genotype landraces. Samples were gathered for each landrace by mixing equal weights from young leaves, from which DNA was extracted. We then estimated allelic frequencies in each DNA bulk based on fluorescent intensity ratio (FIR) between two alleles at each SNP using a two step-approach. We first tested either whether the DNA bulk was monomorphic or polymorphic according to the two FIR distributions of individuals homozygous for allele A or B, respectively. If the DNA bulk was polymorphic, we estimated its allelic frequency by using a predictive equation calibrated on FIR from DNA bulks with known allelic frequencies. Our approach: (i) gives accurate allelic frequency estimations that are highly reproducible across laboratories, (ii) protects against false detection of allele fixation within landraces. We estimated allelic frequencies of 23,412 SNPs in 156 landraces representing American and European maize diversity. Modified Roger’s genetic Distance between 156 landraces estimated from 23,412 SNPs and 17 SSRs using the same DNA bulks were highly correlated, suggesting that the ascertainment bias is low. Our approach is affordable, easy to implement and does not require specific bioinformatics support and laboratory equipment, and therefore should be highly relevant for large-scale characterization of genebanks for a wide range of species.

scholarly journals Deciphering the Genetic Diversity of Landraces With High-Throughput SNP Genotyping of DNA Bulks: Methodology and Application to the Maize 50k Array

2021 ◽  
Vol 11 ◽  
Author(s):  
Mariangela Arca ◽  
Tristan Mary-Huard ◽  
Brigitte Gouesnard ◽  
Aurélie Bérard ◽  
Cyril Bauland ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Throughput ◽  
Large Scale ◽  
Allelic Frequency ◽  
Laboratory Equipment ◽  
Fluorescent Intensity ◽  
Allelic Frequencies ◽  
Wide Range ◽  
Young Leaves ◽  
Scale Characterization

Genebanks harbor original landraces carrying many original favorable alleles for mitigating biotic and abiotic stresses. Their genetic diversity remains, however, poorly characterized due to their large within genetic diversity. We developed a high-throughput, cheap and labor saving DNA bulk approach based on single-nucleotide polymorphism (SNP) Illumina Infinium HD array to genotype landraces. Samples were gathered for each landrace by mixing equal weights from young leaves, from which DNA was extracted. We then estimated allelic frequencies in each DNA bulk based on fluorescent intensity ratio (FIR) between two alleles at each SNP using a two step-approach. We first tested either whether the DNA bulk was monomorphic or polymorphic according to the two FIR distributions of individuals homozygous for allele A or B, respectively. If the DNA bulk was polymorphic, we estimated its allelic frequency by using a predictive equation calibrated on FIR from DNA bulks with known allelic frequencies. Our approach: (i) gives accurate allelic frequency estimations that are highly reproducible across laboratories, (ii) protects against false detection of allele fixation within landraces. We estimated allelic frequencies of 23,412 SNPs in 156 landraces representing American and European maize diversity. Modified Roger’s genetic Distance between 156 landraces estimated from 23,412 SNPs and 17 simple sequence repeats using the same DNA bulks were highly correlated, suggesting that the ascertainment bias is low. Our approach is affordable, easy to implement and does not require specific bioinformatics support and laboratory equipment, and therefore should be highly relevant for large-scale characterization of genebanks for a wide range of species.

scholarly journals Transcriptome wide SSR discovery cross-taxa transferability and development of marker database for studying genetic diversity population structure of Lilium species

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Manosh Kumar Biswas ◽  
Mita Bagchi ◽  
Ujjal Kumar Nath ◽  
Dhiman Biswas ◽  
Sathishkumar Natarajan ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Population Structure ◽  
Large Scale ◽  
Polymorphic Information Content ◽  
Pcr Amplification ◽  
Efficient Manner ◽  
Lilium Species ◽  
Population Structure Analysis ◽  
Wide Range ◽  
Functional Molecular Markers

Abstract Lily belongs to family liliaceae, which mainly propagates vegetatively. Therefore, sufficient number of polymorphic, informative, and functional molecular markers are essential for studying a wide range of genetic parameters in Lilium species. We attempted to develop, characterize and design SSR (simple sequence repeat) markers using online genetic resources for analyzing genetic diversity and population structure of Lilium species. We found di-nucleotide repeat motif were more frequent (4684) within 0.14 gb (giga bases) transcriptome than other repeats, of which was two times higher than tetra-repeat motifs. Frequency of di-(AG/CT), tri-(AGG/CTT), tetra-(AAAT), penta-(AGAGG), and hexa-(AGAGGG) repeats was 34.9%, 7.0%, 0.4%, 0.3%, and 0.2%, respectively. A total of 3607 non-redundant SSR primer pairs was designed based on the sequences of CDS, 5′-UTR and 3′-UTR region covering 34%, 14%, 23%, respectively. Among them, a sub set of primers (245 SSR) was validated using polymerase chain reaction (PCR) amplification, of which 167 primers gave expected PCR amplicon and 101 primers showed polymorphism. Each locus contained 2 to 12 alleles on average 0.82 PIC (polymorphic information content) value. A total of 87 lily accessions was subjected to genetic diversity analysis using polymorphic SSRs and found to separate into seven groups with 0.73 to 0.79 heterozygosity. Our data on large scale SSR based genetic diversity and population structure analysis may help to accelerate the breeding programs of lily through utilizing different genomes, understanding genetics and characterizing germplasm with efficient manner.

scholarly journals Automation assisted anaerobic phenotyping for metabolic engineering

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Kaushik Raj ◽  
Naveen Venayak ◽  
Patrick Diep ◽  
Sai Akhil Golla ◽  
Alexander F. Yakunin ◽  
...  
Keyword(s):  
Metabolic Engineering ◽  
High Throughput ◽  
Large Scale ◽  
Plastic Waste ◽  
Small Scale ◽  
Chemical Production ◽  
Liquid Handling ◽  
Strain Design ◽  
Wide Range ◽  
Scale Down

Abstract Background Microorganisms can be metabolically engineered to produce a wide range of commercially important chemicals. Advancements in computational strategies for strain design and synthetic biological techniques to construct the designed strains have facilitated the generation of large libraries of potential candidates for chemical production. Consequently, there is a need for high-throughput laboratory scale techniques to characterize and screen these candidates to select strains for further investigation in large scale fermentation processes. Several small-scale fermentation techniques, in conjunction with laboratory automation have enhanced the throughput of enzyme and strain phenotyping experiments. However, such high throughput experimentation typically entails large operational costs and generate massive amounts of laboratory plastic waste. Results In this work, we develop an eco-friendly automation workflow that effectively calibrates and decontaminates fixed-tip liquid handling systems to reduce tip waste. We also investigate inexpensive methods to establish anaerobic conditions in microplates for high-throughput anaerobic phenotyping. To validate our phenotyping platform, we perform two case studies—an anaerobic enzyme screen, and a microbial phenotypic screen. We used our automation platform to investigate conditions under which several strains of E. coli exhibit the same phenotypes in 0.5 L bioreactors and in our scaled-down fermentation platform. We also propose the use of dimensionality reduction through t-distributed stochastic neighbours embedding (t-SNE) in conjunction with our phenotyping platform to effectively cluster similarly performing strains at the bioreactor scale. Conclusions Fixed-tip liquid handling systems can significantly reduce the amount of plastic waste generated in biological laboratories and our decontamination and calibration protocols could facilitate the widespread adoption of such systems. Further, the use of t-SNE in conjunction with our automation platform could serve as an effective scale-down model for bioreactor fermentations. Finally, by integrating an in-house data-analysis pipeline, we were able to accelerate the ‘test’ phase of the design-build-test-learn cycle of metabolic engineering.

scholarly journals Aging Atlas: a multi-omics database for aging biology

2020 ◽  
Vol 49 (D1) ◽  
pp. D825-D830 ◽  
Author(s):  
◽  
Guang-Hui Liu ◽  
Yiming Bao ◽  
Jing Qu ◽  
Weiqi Zhang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
High Throughput ◽  
Large Scale ◽  
Wide Spectrum ◽  
Molecular Profile ◽  
Aging Research ◽  
Related Data ◽  
Age Related ◽  
Wide Range

Abstract Organismal aging is driven by interconnected molecular changes encompassing internal and extracellular factors. Combinational analysis of high-throughput ‘multi-omics’ datasets (gathering information from genomics, epigenomics, transcriptomics, proteomics, metabolomics and pharmacogenomics), at either populational or single-cell levels, can provide a multi-dimensional, integrated profile of the heterogeneous aging process with unprecedented throughput and detail. These new strategies allow for the exploration of the molecular profile and regulatory status of gene expression during aging, and in turn, facilitate the development of new aging interventions. With a continually growing volume of valuable aging-related data, it is necessary to establish an open and integrated database to support a wide spectrum of aging research. The Aging Atlas database aims to provide a wide range of life science researchers with valuable resources that allow access to a large-scale of gene expression and regulation datasets created by various high-throughput omics technologies. The current implementation includes five modules: transcriptomics (RNA-seq), single-cell transcriptomics (scRNA-seq), epigenomics (ChIP-seq), proteomics (protein–protein interaction), and pharmacogenomics (geroprotective compounds). Aging Atlas provides user-friendly functionalities to explore age-related changes in gene expression, as well as raw data download services. Aging Atlas is freely available at https://bigd.big.ac.cn/aging/index.

scholarly journals Automation assisted anaerobic phenotyping for metabolic engineering

2021 ◽  
Author(s):  
Kaushik Raj ◽  
Naveen Venayak ◽  
Patrick Diep ◽  
Sai Akhil Golla ◽  
Alexander F. Yakunin ◽  
...  
Keyword(s):  
Metabolic Engineering ◽  
High Throughput ◽  
Large Scale ◽  
Small Scale ◽  
Chemical Production ◽  
E Coli ◽  
Strain Design ◽  
Phenotypic Screen ◽  
Wide Range ◽  
Scale Down

Microorganisms can be metabolically engineered to produce a wide range of commercially important chemicals. Advancements in computational strategies for strain design and synthetic biological techniques to construct the designed strains have facilitated the generation of large libraries of potential candidates for chemical production. Consequently, there is a need for a high-throughput, laboratory scale methods to characterize and screen these candidates to select strains for further investigation in large scale fermentation processes. Several small-scale fermentation techniques, in conjunction with laboratory automation have enhanced the throughput of enzyme and strain phenotyping experiments. However, such high throughput experimentation typically entails large operational costs and generate massive amounts of laboratory plastic waste. In this work, we develop an eco-friendly automation workflow that effectively calibrates and decontaminates fixed-tip liquid handling systems to reduce tip waste. We also investigate inexpensive methods to establish anaerobic conditions in microplates for high-throughput anaerobic phenotyping. To validate our phenotyping platform, we assess its performance in two case studies - an anaerobic enzyme screen, and a microbial phenotypic screen. We used our automation platform to investigate conditions under which several strains of E. coli exhibit the same phenotypes in 0.5 L bioreactors and in our scaled-down fermentation platform. Further, we propose the use of dimensionality reduction through t-distributed stochastic neighbours embedding, in conjunction with our phenotyping platform to serve as an effective scale-down model for bioreactor phenotypes. By integrating an in-house data-analysis pipeline, we were able to accelerate the 'test' phase of the design-build-test-learn cycle of metabolic engineering.

scholarly journals The changing landscape of astrostatistics and astroinformatics

2016 ◽  
Vol 12 (S325) ◽  
pp. 3-9
Author(s):  
Eric D. Feigelson
Keyword(s):  
Statistical Methods ◽  
High Throughput ◽  
Data Reduction ◽  
Large Scale ◽  
Formal Education ◽  
Current Status ◽  
Statistical Computing ◽  
Software Environment ◽  
Wide Range ◽  
Computational Methodology

AbstractThe history and current status of the cross-disciplinary fields of astrostatistics and astroinformatics are reviewed. Astronomers need a wide range of statistical methods for both data reduction and science analysis. With the proliferation of high-throughput telescopes, efficient large scale computational methods are also becoming essential. However, astronomers receive only weak training in these fields during their formal education. Interest in the fields is rapidly growing with conferences organized by scholarly societies, textbooks and tutorial workshops, and research studies pushing the frontiers of methodology. R, the premier language of statistical computing, can provide an important software environment for the incorporation of advanced statistical and computational methodology into the astronomical community.

scholarly journals Characterisation of a Novel Benzopyran Library Using High-Throughput Microscopy

2021 ◽  
Author(s):  
◽  
Jeffrey Sheridan
Keyword(s):  
Functional Groups ◽  
High Throughput ◽  
Large Scale ◽  
Structural Diversity ◽  
Ergosterol Biosynthesis ◽  
Biological Interactions ◽  
Diverse Range ◽  
Genome Wide ◽  
Wide Range ◽  
Disciplinary Field

<p>Drug discovery is a multi-disciplinary field incorporating both chemistry and biology to create novel pharmaceuticals. Nature synthesizes a diverse range of chemical entities that can demonstrate a wide range of biological interactions, though often produces these compounds in small amounts. Using natures structural diversity as a template, organic synthetic chemistry can tap into the structures of natural products and provide novel structures as well as overcome supply issues through large-scale synthetic chemical processes. A novel benzopyran library was synthesised by Sandile Simelane by reacting 3,4,6,-tri-O-acetyl-D-galactal with various phenols to create a novel focused library of bridged benzopyrans. Each molecule has unique functional groups at defined points in the structure due to varying the functional groups on the phenol, allowing for variation within the library whilst retaining the core scaffold. In this thesis, the bioactivity of this novel benzopyran library was explored using a phenotypic screen measuring growth inhibition. A compound, S13, was determined to be the most potent in the library, therefore genome-wide screening was performed using S13. High-throughput microscopy of 4,100 strains, each with a different GFP-tagged protein, was utilized to determine proteins that increased in abundance or changed localization in response to perturbation with S13. Following treatment with S13, the yeast vacuole increased in size due to an aggregation of proteins in the vacuolar lumen. The increase in vacuole size was coincident with a decrease in vacuolar acidity, potentially disrupted autophagy and the upregulation of several proteins involved in ergosterol biosynthesis. Together, these results reveal a novel bridged benzopyran that increases vacuolar size and pH through an epistatic mechanism involving ergosterol biosynthesis.</p>

Advanced technology in the hospital laboratory: Equipment for clinical chemistry and its evaluation

1973 ◽  
Vol 184 (1077) ◽  
pp. 351-359 ◽  
Keyword(s):  
Quality Control ◽  
Large Scale ◽  
Clinical Chemistry ◽  
Advanced Technology ◽  
Laboratory Equipment ◽  
Fast Computer ◽  
Wide Range ◽  
Computer Controlled ◽  
Effectiveness And Efficiency ◽  
The Right

The last 20 years have seen the introduction of automation into clinical chemistry on a large scale and much of the once familiar equipment - burettes, pipettes, test tubes etc., have largely disappeared. Manual error can be minimized and complex procedures for quality control successfully applied. A wide range of instruments are now available from simple dispensers and diluters to complex, fast, computer controlled twenty channel analysers producing twenty results every 12 s. The value of the most expensive item of equipment in the laboratory has increased from £30 to £100 000 and it is important that expensive items are fully evaluated for cost effectiveness and efficiency so that the right purchase is made for each situation.

scholarly journals Characterisation of a Novel Benzopyran Library Using High-Throughput Microscopy

2021 ◽  
Author(s):  
◽  
Jeffrey Sheridan
Keyword(s):  
Functional Groups ◽  
High Throughput ◽  
Large Scale ◽  
Structural Diversity ◽  
Ergosterol Biosynthesis ◽  
Biological Interactions ◽  
Diverse Range ◽  
Genome Wide ◽  
Wide Range ◽  
Disciplinary Field

<p>Drug discovery is a multi-disciplinary field incorporating both chemistry and biology to create novel pharmaceuticals. Nature synthesizes a diverse range of chemical entities that can demonstrate a wide range of biological interactions, though often produces these compounds in small amounts. Using natures structural diversity as a template, organic synthetic chemistry can tap into the structures of natural products and provide novel structures as well as overcome supply issues through large-scale synthetic chemical processes. A novel benzopyran library was synthesised by Sandile Simelane by reacting 3,4,6,-tri-O-acetyl-D-galactal with various phenols to create a novel focused library of bridged benzopyrans. Each molecule has unique functional groups at defined points in the structure due to varying the functional groups on the phenol, allowing for variation within the library whilst retaining the core scaffold. In this thesis, the bioactivity of this novel benzopyran library was explored using a phenotypic screen measuring growth inhibition. A compound, S13, was determined to be the most potent in the library, therefore genome-wide screening was performed using S13. High-throughput microscopy of 4,100 strains, each with a different GFP-tagged protein, was utilized to determine proteins that increased in abundance or changed localization in response to perturbation with S13. Following treatment with S13, the yeast vacuole increased in size due to an aggregation of proteins in the vacuolar lumen. The increase in vacuole size was coincident with a decrease in vacuolar acidity, potentially disrupted autophagy and the upregulation of several proteins involved in ergosterol biosynthesis. Together, these results reveal a novel bridged benzopyran that increases vacuolar size and pH through an epistatic mechanism involving ergosterol biosynthesis.</p>

scholarly journals A multispecies amplicon sequencing approach for genetic diversity assessment in grassland plant species

2021 ◽  
Author(s):  
Miguel Loera-Sanchez ◽  
Bruno Studer ◽  
Roland Koelliker
Keyword(s):  
Genetic Diversity ◽  
Plant Species ◽  
High Throughput ◽  
Large Scale ◽  
Trifolium Pratense ◽  
Amplicon Sequencing ◽  
Pathogen Resistance ◽  
Production Plant ◽  
Diversity Assessment ◽  
Grassland Plants

Grasslands are widespread and economically relevant ecosystems at the basis of sustainable roughage production. Plant genetic diversity (PGD; i.e., within-species diversity) is related to many beneficial effects to the ecosystem functioning of grasslands. The monitoring of PGD in temperate grasslands is complicated by the multiplicity of species present and by a shortage of methods for large-scale assessment. However, the continuous advancement of high-throughput DNA sequencing approaches have improved the prospects of broad, multispecies PGD monitoring. Among them, amplicon sequencing stands out as a robust and cost-effective method. Here we report a set of twelve multispecies primer pairs that can be used for high-throughput PGD assessment in multiple grassland plant species. The loci targeted by the amplicons were selected and tested in two phases: a "discovery phase" based on a sequence capture assay (611 target nuclear loci assessed in 16 grassland plant species), which resulted in the selection of eleven loci; and a "validation phase", in which the selected loci were targeted and sequenced using twelve multispecies primers in test populations of Dactylis glomerata L., Lolium perenne L., Festuca pratensis Huds., Trifolium pratense L. and T. repens L. The resulting multispecies amplicons had overall nucleotide diversities per species ranging from 5.19 × 10-3 to 1.29 × 10-2, which is in the range of flowering-related genes but slightly lower than pathogen resistance genes. We conclude that the methodology, the DNA sequence resources, and the amplicon-specific primer pairs reported in this study provide the basis for large-scale, multispecies PGD monitoring in grassland plants.

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