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Constructed Wetlands Technology in Cuba: Research Experiences
Published in María del Carmen Durán-Domínguez-de-Bazúa, Amado Enrique Navarro-Frómeta, Josep M. Bayona, Artificial or Constructed Wetlands, 2018
Irina Salgado-Bernal, Maira M. Perez-Villar, Lizandra Perez-Bou, Mario Cruz-Arias, Margie Zorrilla-Velazco, Maria E. Carballo-Valdes
Metagenomics can be a useful tool for this purpose, because it allows us to know how many and what species are present in a certain atmosphere and what ecological function play each species in an individual way. The term metagenomics was coined by Handelsman et al. in 1998. Among the concepts defined in the literature, one of the most synthetic belongs to Chen and Pachter (2005), that defines it as the application of modern genomic techniques for the study of microbial communities directly in their natural atmospheres. Other authors like Riesenfeld et al. (2004) define it as the investigation of collective microbial genomes retrieved directly from environmental samples and does not rely on cultivation or prior knowledge of the microbial communities. Traditional cultivation methods and traditional genomics can at best access 1%. However, metagenomics can in principle access 100% of the genetic resources in an environment. DNA is directly extracted from the environment and cloned into cosmid, fosmid, or bacterial artificial chromosomes (BAC) vectors producing large insert libraries.
Functional Metagenomics for Bioprospecting Novel Glycosyl Hydrolases in 2G Biofuel Production from Lignocellulosics
Published in Jitendra Kumar Saini, Surender Singh, Lata Nain, Sustainable Microbial Technologies for Valorization of Agro-Industrial Wastes, 2023
Sugitha Thankappan, Sajan Kurien, Surender Singh, Asish K. Binodh
In general, the genes associated to interrelated metabolic pathways present as gene cassettes like operons/super-operonic clusters in prokaryotes contrast to eukaryotes. Thus, for functional screening, it is important to clone the metagenomic DNA into suitable vectors, like fosmids or cosmids. Further, promoters of a multiple-cloning site (MCS) help bidirectional transcription, which increases the probability of positive clones with the targets (Lämmle et al., 2007). Further, broad host range systems are required to enhance expression successfully and in the identification of targeted genes, as the gene expression significantly depends on the host (Yoon et al., 2013). Although E. coli is highly economical, and effective, for production of a wide range of heterologous proteins (Hannig & Makrides, 1998), it poses certain limitations in the number of GHs obtained from metagenomic libraries from an enriched environment (L.-L. Li, McCorkle, Monchy, Taghavi, & van der Lelie, 2009). Furthermore, the application of bacterial host systems reduces the chances of identifying GHs of fungal origin, due to specific codon usage, regulation, and activation of promoter, followed by posttranslational process. These factors impact the expression of eukaryotic genes in prokaryotic systems more specifically at their functional level. Likewise, prokaryotic host systems lack some posttranslational events, thereby affecting their production extracellularly (Flipphi et al., 2013; Juturu & Wu, 2014; L.-L. Li et al., 2009). Since majority of the GHs identified so far belong to prokaryotic systems, alternate host expression systems need to be regarded in other genera. T. thermophilus is a successful metagenomic library host in terms of more active clones for screening esterases and expression of xylanases (Angelov, Mientus, Liebl, & Liebl, 2009; Leis et al., 2015). The ongoing projects on the development of eukaryotic host systems like yeast cells (Pichia pastoris) will address the limitations of functional expression of eukaryotic LCB-deconstructing enzymes.
Molecular biological tools in concrete biodeterioration – a mini review
Published in Environmental Technology, 2019
Vinita Vishwakarma, Balakrishnan Anandkumar
The genome analysis gives the details of complete DNA sequences, coding and noncoding part of genome. The analysis is based on the gene identification and then manipulation. In the whole genome sequencing (WGS) approach, the genomes of all microorganisms present in the environmental sample are broken into small fragments and the fragments will be cloned into vectors as plasmid or fosmid gene subclone libraries. The sub clone libraries will be sequenced and the above said ‘shotgun’ sequencing will be done with any one of the following sequencing platforms such as (a) the Roche/454 FLX [86] (http://www.454.com/enablingtechnology/the-system.asp), (b) the Illumina/Solexa Genome Analyzer [87] (http://www.illumina.com/pages.ilmn?ID=203), (c) and the Applied Biosystems SOLiDTM System (http://marketingappliedbiosystems.com/images/Product/SolidKnowledge/flash/102207/solid.html), (d) the Helicos HeliscopeTM (www.helicosbio.com) and (e) Pacific Biosciences SMRT (www.pacificbiosciences.com) instruments [88]. The sequencing reads and the contig assembly will provide the whole genome with the use of assembly algorithms. The complexity of the community and the relative abundance, the key factors for the successful genome assembly will enable to characterize the ecological biodiversity and the identification of unknown aetiological agents [88,89]. The analysis of whole microbial genomes is the way to know the microbial evolution and diversity beyond single protein or gene phylogenies. The whole microbial genomes analysis is also a powerful tool in identifying new bacterial groups involved in MICC and it may pave a platform for controlling the bacterial groups with concrete having the biocides. The whole metagenome of biofilms in a corroded sewer pipe was sequenced and analysed by Gomez-Alvarez et al. [85] shed the information about the microbial composition and functional marker genes associated with the biofilms. An abundant SOB and SRB group members along with the other bacterial phyla could be identified and analysed using this approach [85]. Gomez-Alvarez et al. also demonstrated, combined with transcriptomics, an enrichment of genes associated with heavy metal resistance, virulence (protein secretion systems) and stress response in the biofilm analysed [85]. This whole biofilm genome analysis can be implemented to understand the complete microbial network and functional gene markers to evaluate and mitigate MICC in the biofilms formed on the different concrete structures.
Molecular biological tools in concrete biodeterioration – a mini review
Published in Environmental Technology Reviews, 2018
Vinita Vishwakarma, Balakrishnan Anandkumar
The genome analysis gives the details of complete DNA sequences, coding and noncoding part of genome. The analysis is based on the gene identification and then manipulation. In the whole genome sequencing (WGS) approach, the genomes of all microorganisms present in the environmental sample are broken into small fragments and the fragments will be cloned into vectors as plasmid or fosmid gene subclone libraries. The sub clone libraries will be sequenced and the above said ‘shotgun’ sequencing will be done with any one of the following sequencing platforms such as (a) the Roche/454 FLX [86] (http://www.454.com/enablingtechnology/the-system.asp), (b) the Illumina/Solexa Genome Analyzer [87] (http://www.illumina.com/pages.ilmn?ID=203), (c) and the Applied Biosystems SOLiDTM System (http://marketingappliedbiosystems.com/images/Product/SolidKnowledge/flash/102207/solid.html), (d) the Helicos HeliscopeTM (www.helicosbio.com) and (e) Pacific Biosciences SMRT (www.pacificbiosciences.com) instruments [88]. The sequencing reads and the contig assembly will provide the whole genome with the use of assembly algorithms. The complexity of the community and the relative abundance, the key factors for the successful genome assembly will enable to characterize the ecological biodiversity and the identification of unknown aetiological agents [88,89]. The analysis of whole microbial genomes is the way to know the microbial evolution and diversity beyond single protein or gene phylogenies. The whole microbial genomes analysis is also a powerful tool in identifying new bacterial groups involved in MICC and it may pave a platform for controlling the bacterial groups with concrete having the biocides. The whole metagenome of biofilms in a corroded sewer pipe was sequenced and analysed by Gomez-Alvarez et al. [85] shed the information about the microbial composition and functional marker genes associated with the biofilms. An abundant SOB and SRB group members along with the other bacterial phyla could be identified and analysed using this approach [85]. Gomez-Alvarez et al. also demonstrated, combined with transcriptomics, an enrichment of genes associated with heavy metal resistance, virulence (protein secretion systems) and stress response in the biofilm analysed [85]. This whole biofilm genome analysis can be implemented to understand the complete microbial network and functional gene markers to evaluate and mitigate MICC in the biofilms formed on the different concrete structures.