Construction of Small-Insert and Large-Insert Metagenomic Libraries

Simon, Chantal; Daniel, Rolf

doi:10.1007/978-1-0716-2795-2_1

Cited by 3 publications

(3 citation statements)

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“…The function‐based metagenomics approach involves the construction of metagenomic libraries containing fragments of community DNA and screening for a specific phenotype (Dinsdale et al, 2008). The steps involved are enzymatic/ mechanical shearing of DNA, ligation to a vector, and transformation into a heterologous host (Simon & Daniel, 2017). Two types of libraries may be constructed depending on the size of the insert.…”

Section: The Dawn Of Metagenomics Eramentioning

confidence: 99%

A primer on metagenomics and next‐generation sequencing in fish gut microbiome research

Johny

Puthusseri

Bhat

2021

Aquaculture Research

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Fishes constitute a vital component of the aquatic ecosystem, that interacts with physical, chemical, and biological components of the ecosystem. They are found in almost all aquatic environments ranging from high mountain streams to the abyssal and hadal zones of the deepest oceans. With an origin dating back to 600 million years and an estimate of 34,600 species described till date (FishBase, 2021), fishes comprise nearly half of the total vertebrate diversity (Nelson, 2007).Fishes are associated with complex assemblages of microorganisms, comprising bacteria, archaea, viruses, yeasts, and protists, collectively known as 'microbiota' or 'microbiome' (Merrifield & Rodiles, 2015). The characteristic microbial community inhabiting a particular environment along with their genomes and theatre of activity is referred to as the 'microbiome' (Gilbert et al., 2016;Prescott, 2017). However, in certain contexts, the term 'microbiota' is used to define the collection of microorganisms and the term 'microbiome' to refers to the collection of microbial genomes (Burokas et al., 2015).Microorganisms take residence in almost every organ of the fish such as skin, gills, digestive tract, internal organs like the liver, kidney, spleen, and light-emitting organs (Austin, 2002). However, fish gut microflora is often acclaimed as an 'extra organ' owing to its significant contribution to important physiological functions of the host such as digestion, metabolism, reproduction, development, and immune response (Butt & Volkoff, 2019). Further, the composition of the gut microflora is affected by a plethora of factors such as habitat salinity, temperature, trophic level, taxonomy, feeding habits and host selective pressures within the gut, leading to the development of a unique and diverse microflora (Bevins & Salzman, 2011;Sullam et al., 2012). Therefore, it may be safely presumed that the fish gut microbiome, representing 'worlds within worlds' (Butt & Volkoff, 2019) is a biodiversity hotspot and a promising source for bioprospecting applications.

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Section: The Dawn Of Metagenomics Eramentioning

confidence: 99%

A primer on metagenomics and next‐generation sequencing in fish gut microbiome research

Johny

Puthusseri

Bhat

2021

Aquaculture Research

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“…Short-insert genomic libraries are created by inserting small DNA fragments into plasmid vectors. Large inserts can be cloned onto BAC or cosmid vectors, which can carry inserts larger than 40 Kb, or fosmid or cosmid vectors, which can handle inserts up to 40 Kb in size [12].…”

Section: Introductionmentioning

confidence: 99%

The Role of Metagenomic Approaches in the Analysis of Microbial Community in Extreme Environment

Shuikan¹,

Alshuwaykan²,

Arif³

2023

Life in Extreme Environments - Diversity, Adaptability and Valuable Resources of Bioactive Molecules

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Metagenomic is a promising technique that has many applications in different fields. In fact, metagenomics is the ideal culture-independent technique that unravels the microbial composition and biodiversity in the sample, which helps scientists to study and understand how this biodiversity is affected by continuously changing conditions in the environment and how this microbial community interacts with each other. In the past, the microbial composition in extreme environments was undiscovered due to the difficulty of isolation, culturing, and identification of microbes living there. However, nowadays after the development and combination of metagenomic and next-generation sequencing techniques, it became more easy to study the microbial composition in extreme environments without culturing. In this chapter, the use of metagenomic techniques to study the microbial biodiversity in different extreme environments are discussed. In addition, different NGS platforms are discussed in terms of principles, advantages, and limitations.

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“…Many applications of fundamental importance in modern biotechnology and biomedicine, DNA amplification methods (including polymerase chain reaction (PCR)), and some of the most cutting-edge DNA sequencing technologies would not be possible without advances in the structure and function of DNAPs [1]. Among these techniques, whole genome amplification (WGA) refers to the amplification of genome DNA sequences in a sample that can range from a single virus [2] to polyploid eukaryotic genomes [3] or complex metagenomes [4]. WGA typically starts from a tiny DNA input, providing enough genetic material for subsequent analyses on the order of micrograms.…”

Section: Introductionmentioning

confidence: 99%

DNA Polymerases for Whole Genome Amplification: Considerations and Future Directions

Ordóñez

Redrejo-Rodríguez

2023

IJMS

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In the same way that specialized DNA polymerases (DNAPs) replicate cellular and viral genomes, only a handful of dedicated proteins from various natural origins as well as engineered versions are appropriate for competent exponential amplification of whole genomes and metagenomes (WGA). Different applications have led to the development of diverse protocols, based on various DNAPs. Isothermal WGA is currently widely used due to the high performance of Φ29 DNA polymerase, but PCR-based methods are also available and can provide competent amplification of certain samples. Replication fidelity and processivity must be considered when selecting a suitable enzyme for WGA. However, other properties, such as thermostability, capacity to couple replication, and double helix unwinding, or the ability to maintain DNA replication opposite to damaged bases, are also very relevant for some applications. In this review, we provide an overview of the different properties of DNAPs widely used in WGA and discuss their limitations and future research directions.

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Construction of Small-Insert and Large-Insert Metagenomic Libraries

Cited by 3 publications

References 10 publications

A primer on metagenomics and next‐generation sequencing in fish gut microbiome research

A primer on metagenomics and next‐generation sequencing in fish gut microbiome research

The Role of Metagenomic Approaches in the Analysis of Microbial Community in Extreme Environment

DNA Polymerases for Whole Genome Amplification: Considerations and Future Directions

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