scholarly journals High-Throughput Selection and Characterisation of Aptamers on Optical Next-Generation Sequencers

2021 ◽  
Vol 22 (17) ◽  
pp. 9202
Author(s):  
Alissa Drees ◽  
Markus Fischer
Keyword(s):  
High Throughput ◽  
High Performance ◽  
High Throughput Sequencing ◽  
Selection Process ◽  
Next Generation ◽  
Binding Assays ◽  
Ligand Interaction ◽  
Modified Dna ◽  
Sequencing Platforms ◽  
And Function

Aptamers feature a number of advantages, compared to antibodies. However, their application has been limited so far, mainly because of the complex selection process. ‘High-throughput sequencing fluorescent ligand interaction profiling’ (HiTS–FLIP) significantly increases the selection efficiency and is consequently a very powerful and versatile technology for the selection of high-performance aptamers. It is the first experiment to allow the direct and quantitative measurement of the affinity and specificity of millions of aptamers simultaneously by harnessing the potential of optical next-generation sequencing platforms to perform fluorescence-based binding assays on the clusters displayed on the flow cells and determining their sequence and position in regular high-throughput sequencing. Many variants of the experiment have been developed that allow automation and in situ conversion of DNA clusters into base-modified DNA, RNA, peptides, and even proteins. In addition, the information from mutational assays, performed with HiTS–FLIP, provides deep insights into the relationship between the sequence, structure, and function of aptamers. This enables a detailed understanding of the sequence-specific rules that determine affinity, and thus, supports the evolution of aptamers. Current variants of the HiTS–FLIP experiment and its application in the field of aptamer selection, characterisation, and optimisation are presented in this review.

scholarly journals Assessment of Bioleaching Microbial Community Structure and Function Based on Next-Generation Sequencing Technologies

Minerals ◽  
2018 ◽  
Vol 8 (12) ◽  
pp. 596 ◽  
Author(s):  
Shuang Zhou ◽  
Min Gan ◽  
Jianyu Zhu ◽  
Xinxing Liu ◽  
Guanzhou Qiu
Keyword(s):  
Community Structure ◽  
Next Generation Sequencing ◽  
Microbial Ecology ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Structure And Function ◽  
Next Generation ◽  
Sequencing Technologies ◽  
And Function ◽  
Generation Sequencing

It is widely known that bioleaching microorganisms have to cope with the complex extreme environment in which microbial ecology relating to community structure and function varies across environmental types. However, analyses of microbial ecology of bioleaching bacteria is still a challenge. To address this challenge, numerous technologies have been developed. In recent years, high-throughput sequencing technologies enabling comprehensive sequencing analysis of cellular RNA and DNA within the reach of most laboratories have been added to the toolbox of microbial ecology. The next-generation sequencing technology allowing processing DNA sequences can produce available draft genomic sequences of more bioleaching bacteria, which provides the opportunity to predict models of genetic and metabolic potential of bioleaching bacteria and ultimately deepens our understanding of bioleaching microorganism. High-throughput sequencing that focuses on targeted phylogenetic marker 16S rRNA has been effectively applied to characterize the community diversity in an ore leaching environment. RNA-seq, another application of high-throughput sequencing to profile RNA, can be for both mapping and quantifying transcriptome and has demonstrated a high efficiency in quantifying the changing expression level of each transcript under different conditions. It has been demonstrated as a powerful tool for dissecting the relationship between genotype and phenotype, leading to interpreting functional elements of the genome and revealing molecular mechanisms of adaption. This review aims to describe the high-throughput sequencing approach for bioleaching environmental microorganisms, particularly focusing on its application associated with challenges.

scholarly journals Behavior Individuality: A Focus on Drosophila melanogaster

2021 ◽  
Vol 12 ◽  
Author(s):  
Rubén Mollá-Albaladejo ◽  
Juan A. Sánchez-Alcañiz
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Great Effort ◽  
Behavioral Differences ◽  
Adult Behavior ◽  
Large Numbers ◽  
Environmental Variations ◽  
And Function

Among individuals, behavioral differences result from the well-known interplay of nature and nurture. Minute differences in the genetic code can lead to differential gene expression and function, dramatically affecting developmental processes and adult behavior. Environmental factors, epigenetic modifications, and gene expression and function are responsible for generating stochastic behaviors. In the last decade, the advent of high-throughput sequencing has facilitated studying the genetic basis of behavior and individuality. We can now study the genomes of multiple individuals and infer which genetic variations might be responsible for the observed behavior. In addition, the development of high-throughput behavioral paradigms, where multiple isogenic animals can be analyzed in various environmental conditions, has again facilitated the study of the influence of genetic and environmental variations in animal personality. Mainly, Drosophila melanogaster has been the focus of a great effort to understand how inter-individual behavioral differences emerge. The possibility of using large numbers of animals, isogenic populations, and the possibility of modifying neuronal function has made it an ideal model to search for the origins of individuality. In the present review, we will focus on the recent findings that try to shed light on the emergence of individuality with a particular interest in D. melanogaster.

scholarly journals Hi-C chromosome conformation capture sequencing of avian genomes using the BGISEQ-500 platform

GigaScience ◽  
2020 ◽  
Vol 9 (8) ◽  
Author(s):  
Marcela Sandoval-Velasco ◽  
Juan Antonio Rodríguez ◽  
Cynthia Perez Estrada ◽  
Guojie Zhang ◽  
Erez Lieberman Aiden ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
High Throughput Sequencing ◽  
Data Generation ◽  
Next Generation ◽  
Sequencing Data ◽  
Yield Data ◽  
Chromosome Conformation ◽  
Sequencing Platform ◽  
Sequencing Platforms ◽  
Generation Sequencing

Abstract Background Hi-C experiments couple DNA-DNA proximity with next-generation sequencing to yield an unbiased description of genome-wide interactions. Previous methods describing Hi-C experiments have focused on the industry-standard Illumina sequencing. With new next-generation sequencing platforms such as BGISEQ-500 becoming more widely available, protocol adaptations to fit platform-specific requirements are useful to give increased choice to researchers who routinely generate sequencing data. Results We describe an in situ Hi-C protocol adapted to be compatible with the BGISEQ-500 high-throughput sequencing platform. Using zebra finch (Taeniopygia guttata) as a biological sample, we demonstrate how Hi-C libraries can be constructed to generate informative data using the BGISEQ-500 platform, following circularization and DNA nanoball generation. Our protocol is a modification of an Illumina-compatible method, based around blunt-end ligations in library construction, using un-barcoded, distally overhanging double-stranded adapters, followed by amplification using indexed primers. The resulting libraries are ready for circularization and subsequent sequencing on the BGISEQ series of platforms and yield data similar to what can be expected using Illumina-compatible approaches. Conclusions Our straightforward modification to an Illumina-compatible in situHi-C protocol enables data generation on the BGISEQ series of platforms, thus expanding the options available for researchers who wish to utilize the powerful Hi-C techniques in their research.

scholarly journals Technological advancements and their importance for nematode identification

SOIL ◽  
2016 ◽  
Vol 2 (2) ◽  
pp. 257-270 ◽  
Author(s):  
Mohammed Ahmed ◽  
Melanie Sapp ◽  
Thomas Prior ◽  
Gerrit Karssen ◽  
Matthew Alan Back
Keyword(s):  
High Throughput ◽  
Crop Production ◽  
Large Scale ◽  
High Throughput Sequencing ◽  
Biological Indicators ◽  
Rapid Identification ◽  
Community Studies ◽  
Terrestrial Environments ◽  
Traditional Taxonomy ◽  
Sequencing Platforms

Abstract. Nematodes represent a species-rich and morphologically diverse group of metazoans known to inhabit both aquatic and terrestrial environments. Their role as biological indicators and as key players in nutrient cycling has been well documented. Some plant-parasitic species are also known to cause significant losses to crop production. In spite of this, there still exists a huge gap in our knowledge of their diversity due to the enormity of time and expertise often involved in characterising species using phenotypic features. Molecular methodology provides useful means of complementing the limited number of reliable diagnostic characters available for morphology-based identification. We discuss herein some of the limitations of traditional taxonomy and how molecular methodologies, especially the use of high-throughput sequencing, have assisted in carrying out large-scale nematode community studies and characterisation of phytonematodes through rapid identification of multiple taxa. We also provide brief descriptions of some the current and almost-outdated high-throughput sequencing platforms and their applications in both plant nematology and soil ecology.

Performance comparison of benchtop high-throughput sequencing platforms

2012 ◽  
Vol 30 (5) ◽  
pp. 434-439 ◽  
Author(s):  
Nicholas J Loman ◽  
Raju V Misra ◽  
Timothy J Dallman ◽  
Chrystala Constantinidou ◽  
Saheer E Gharbia ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Performance Comparison ◽  
Sequencing Platforms

High-Performance Computing In High-Throughput Sequencing

2013 ◽  
pp. 981-1002 ◽  
Author(s):  
Kamer Kaya ◽  
Ayat Hatem ◽  
Hatice Gülçin Özer ◽  
Kun Huang ◽  
Ümit V. Çatalyürek
Keyword(s):  
High Performance Computing ◽  
High Throughput ◽  
High Performance ◽  
High Throughput Sequencing ◽  
Performance Computing

Screening and Identification of PLK1-Polo Box Binding Peptides by High-Throughput Sequencing of Phage-Selected Libraries

2019 ◽  
Vol 26 (8) ◽  
pp. 620-633 ◽  
Author(s):  
Nousheen Bibi ◽  
Hafsa Niaz ◽  
Ted Hupp ◽  
Mohammad Amjad Kamal ◽  
Sajid Rashid
Keyword(s):  
Next Generation Sequencing ◽  
Phage Display ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
B Lymphocyte ◽  
Fluorescent Protein ◽  
Next Generation ◽  
Peptide Phage Display ◽  
Binding Peptides ◽  
Generation Sequencing

Background: Human proteome contains a plethora of short linear peptide motifs that is crucial for signaling and other cellular processes. These motifs are difficult to identify due to lack of systematic approach for their detection. Objective: Here we demonstrate the use of peptide phage display in combination with high throughput next generation sequencing to identify enriched peptide sequences through biopanning process against polo box domain (PBD) of mitotic polo like kinase 1 (Plk1). Methods: Purified recombinant Plk1 and two unrelated controls namely B-lymphocyte antigen (CD20) and fluorescent protein (mCherry) were subjected to peptide phage display analysis. Bacterially-propagated phage DNA was amplified by PCR using triplet bar coded primers to tag the pool from each amplicon. Results: Proteomic peptide phage display along with next generation sequencing and Bioinformatics analysis demonstrated several known and putative novel interactions which were potentially related to Plk1-PBD. With our strategy, we were able to identify and characterize several Plk1-PBD binding peptides, as well as define more precisely, consensus sequences. Conclusion: We believe that this information could provide valuable tools for exploring novel interaction involved in Plk1 signaling as well as to choose peptides for Plk1 specific drug development.

scholarly journals Implementation of High-Throughput Sequencing (HTS) in Aptamer Selection Technology

2020 ◽  
Vol 21 (22) ◽  
pp. 8774
Author(s):  
Natalia Komarova ◽  
Daria Barkova ◽  
Alexander Kuznetsov
Keyword(s):  
Nucleic Acid ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Combinatorial Libraries ◽  
Structure And Function ◽  
Huge Number ◽  
Systematic Evolution ◽  
And Function ◽  
Exponential Enrichment

Aptamers are nucleic acid ligands that bind specifically to a target of interest. Aptamers have gained in popularity due to their high potential for different applications in analysis, diagnostics, and therapeutics. The procedure called systematic evolution of ligands by exponential enrichment (SELEX) is used for aptamer isolation from large nucleic acid combinatorial libraries. The huge number of unique sequences implemented in the in vitro evolution in the SELEX process imposes the necessity of performing extensive sequencing of the selected nucleic acid pools. High-throughput sequencing (HTS) meets this demand of SELEX. Analysis of the data obtained from sequencing of the libraries produced during and after aptamer isolation provides an informative basis for precise aptamer identification and for examining the structure and function of nucleic acid ligands. This review discusses the technical aspects and the potential of the integration of HTS with SELEX.

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