Construction of Metagenomic DNA Libraries and Enrichment Strategies

2018 ◽  
pp. 23-42
Author(s):  
Farah Fadwa Benbelgacem ◽  
Hamzah Mohd. Salleh ◽  
Ibrahim Ali Noorbatcha
Keyword(s):  
Metagenomic Dna ◽  
Dna Libraries

Construction and validation of two metagenomic DNA libraries from Cerrado soil with high clay content

2011 ◽  
Vol 33 (11) ◽  
pp. 2169-2175 ◽  
Author(s):  
Alinne Pereira de Castro ◽  
Betania Ferraz Quirino ◽  
Heather Allen ◽  
Lynn L. Williamson ◽  
Jo Handelsman ◽  
...  
Keyword(s):  
Clay Content ◽  
Metagenomic Dna ◽  
Dna Libraries

scholarly journals Metagenomic Analyses: Past and Future Trends

2010 ◽  
Vol 77 (4) ◽  
pp. 1153-1161 ◽  
Author(s):  
Carola Simon ◽  
Rolf Daniel
Keyword(s):  
Microbial Communities ◽  
Sequence Data ◽  
Extreme Environments ◽  
Data Sets ◽  
Metagenomic Dna ◽  
Functional Dynamics ◽  
Dna Libraries ◽  
Water Analyses ◽  
Independent Assessment ◽  
Generation Sequencing

ABSTRACTMetagenomics has revolutionized microbiology by paving the way for a cultivation-independent assessment and exploitation of microbial communities present in complex ecosystems. Metagenomics comprising construction and screening of metagenomic DNA libraries has proven to be a powerful tool to isolate new enzymes and drugs of industrial importance. So far, the majority of the metagenomically exploited habitats comprised temperate environments, such as soil and marine environments. Recently, metagenomes of extreme environments have also been used as sources of novel biocatalysts. The employment of next-generation sequencing techniques for metagenomics resulted in the generation of large sequence data sets derived from various environments, such as soil, the human body, and ocean water. Analyses of these data sets opened a window into the enormous taxonomic and functional diversity of environmental microbial communities. To assess the functional dynamics of microbial communities, metatranscriptomics and metaproteomics have been developed. The combination of DNA-based, mRNA-based, and protein-based analyses of microbial communities present in different environments is a way to elucidate the compositions, functions, and interactions of microbial communities and to link these to environmental processes.

scholarly journals Heterologous Expression of the Oxytetracycline Biosynthetic Pathway in Myxococcus xanthus

2010 ◽  
Vol 76 (8) ◽  
pp. 2681-2683 ◽  
Author(s):  
D. Cole Stevens ◽  
Michael R. Henry ◽  
Kimberly A. Murphy ◽  
Christopher N. Boddy
Keyword(s):  
Drug Discovery ◽  
Heterologous Expression ◽  
Biosynthetic Pathway ◽  
Myxococcus Xanthus ◽  
Gene Clusters ◽  
Biosynthetic Pathways ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Metagenomic Dna ◽  
Dna Libraries

ABSTRACT New natural products for drug discovery may be accessed by heterologous expression of bacterial biosynthetic pathways in metagenomic DNA libraries. However, a “universal” host is needed for this experiment. Herein, we show that Myxococcus xanthus is a potential “universal” host for heterologous expression of polyketide biosynthetic gene clusters.

scholarly journals Construction of Aerobic/Anaerobic-Substrate-Induced Gene Expression Procedure for Exploration of Metagenomes From Subseafloor Sediments

2022 ◽  
Vol 12 ◽  
Author(s):  
Taisuke Wakamatsu ◽  
Saki Mizobuchi ◽  
Fumiaki Mori ◽  
Taiki Futagami ◽  
Takeshi Terada ◽  
...  
Keyword(s):  
Gene Expression ◽  
Aromatic Compounds ◽  
Fine Tuning ◽  
Anaerobic Conditions ◽  
Metagenomic Dna ◽  
Library Construction ◽  
Dna Library ◽  
Dna Libraries ◽  
Aerobic And Anaerobic ◽  
Aerobic And Anaerobic Conditions

Substrate-induced gene expression (SIGEX) is a high-throughput promoter-trap method. It is a function-based metagenomic screening tool that relies on transcriptional activation of a reporter gene green fluorescence protein (gfp) by a metagenomic DNA library upon induction with a substrate. However, its use is limited because of the relatively small size of metagenomic DNA libraries and incompatibility with screening metagenomes from anaerobic environments. In this study, these limitations of SIGEX were addressed by fine-tuning metagenome DNA library construction protocol and by using Evoglow, a green fluorescent protein that forms a chromophore even under anaerobic conditions. Two metagenomic libraries were constructed for subseafloor sediments offshore Shimokita Peninsula (Pacific Ocean) and offshore Joetsu (Japan Sea). The library construction protocol was improved by (a) eliminating short DNA fragments, (b) applying topoisomerase-based high-efficiency ligation, (c) optimizing insert DNA concentration, and (d) column-based DNA enrichment. This led to a successful construction of metagenome DNA libraries of approximately 6 Gbp for both samples. SIGEX screening using five aromatic compounds (benzoate, 3-chlorobenzoate, 3-hydroxybenzoate, phenol, and 2,4-dichlorophenol) under aerobic and anaerobic conditions revealed significant differences in the inducible clone ratios under these conditions. 3-Chlorobenzoate and 2,4-dichlorophenol led to a higher induction ratio than that for the other non-chlorinated aromatic compounds under both aerobic and anaerobic conditions. After the further screening of induced clones, a clone induced by 3-chlorobenzoate only under anaerobic conditions was isolated and characterized. The clone harbors a DNA insert that encodes putative open reading frames of unknown function. Previous aerobic SIGEX attempts succeeded in the isolation of gene fragments from anaerobes. This study demonstrated that some gene fragments require a strict in vivo reducing environment to function and may be potentially missed when screened by aerobic induction. The newly developed anaerobic SIGEX scheme will facilitate functional exploration of metagenomes from the anaerobic biosphere.

scholarly journals Construction and validation of metagenomic DNA libraries from landfarm soil microorganisms

2013 ◽  
Vol 12 (2) ◽  
pp. 2148-2155 ◽  
Author(s):  
T.B.A. Pessoa ◽  
S.S. de Souza ◽  
A.F. Cerqueira ◽  
R.P. Rezende ◽  
C.P. Pirovani ◽  
...  
Keyword(s):  
Soil Microorganisms ◽  
Metagenomic Dna ◽  
Dna Libraries

scholarly journals Complementary DNA Libraries and Neurological Disease

1998 ◽  
Vol 55 (6) ◽  
pp. 785
Author(s):  
Benjamin Joseph ◽  
Henry Furneaux
Keyword(s):  
Neurological Disease ◽  
Complementary Dna ◽  
Dna Libraries

Cloning of DNA libraries from mouse Y chromosomes purified by flow cytometry

1986 ◽  
Vol 12 (3) ◽  
pp. 289-295 ◽  
Author(s):  
Bruno Baron ◽  
Philippe Métézeau ◽  
Didier Hatat ◽  
Christa Roberts ◽  
Michel E. Goldberg ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Y Chromosomes ◽  
Dna Libraries
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