Abstract：
:Microbes are now well recognized as major drivers of the biogeochemical cycling that fuels the Earth, and their viruses (phages) are known to be abundant and important in microbial mortality, horizontal gene transfer, and modulating microbial metabolic output. Investigation of environmental phages has been frustrated by an inability to culture the vast majority of naturally occurring diversity coupled with the lack of robust, quantitative, culture-independent methods for studying this uncultured majority. However, for double-stranded DNA phages, a quantitative viral metagenomic sample-to-sequence workflow now exists. Here, we review these advances with special emphasis on the technical details of preparing DNA sequencing libraries for metagenomic sequencing from environmentally relevant low-input DNA samples. Library preparation steps broadly involve manipulating the sample DNA by fragmentation, end repair and adaptor ligation, size fractionation, and amplification. One critical area of future research and development is parallel advances for alternate nucleic acid types such as single-stranded DNA and RNA viruses that are also abundant in nature. Combinations of recent advances in fragmentation (e.g., acoustic shearing and tagmentation), ligation reactions (adaptor-to-template ratio reference table availability), size fractionation (non-gel-sizing), and amplification (linear amplification for deep sequencing and linker amplification protocols) enhance our ability to generate quantitatively representative metagenomic datasets from low-input DNA samples. Such datasets are already providing new insights into the role of viruses in marine systems and will continue to do so as new environments are explored and synergies and paradigms emerge from large-scale comparative analyses.
journal_name
Methods Enzymoljournal_title
Methods in enzymologyauthors
Solonenko SA,Sullivan MBdoi
10.1016/B978-0-12-407863-5.00008-3subject
Has Abstractpub_date
2013-01-01 00:00:00pages
143-65eissn
0076-6879issn
1557-7988pii
B978-0-12-407863-5.00008-3journal_volume
531pub_type
杂志文章abstract：:Genomes can be viewed in terms of their gene content and the order in which the genes appear along each chromosome. Evolutionary events that affect the gene order or content are "rare genomic events" (rarer than events that affect the composition of the nucleotide sequences) and have been advocated by systematists for...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)95035-0
更新日期：2005-01-01 00:00:00
abstract：:ClpB belongs to the Hsp100 family of ring-forming heat-shock proteins involved in degradation of unfolded/misfolded proteins and in reactivation of protein aggregates. ClpB monomers reversibly associate to form the hexameric molecular chaperone that, together with the DnaK system, has the ability to disaggregate stres...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2015.04.007
更新日期：2015-01-01 00:00:00
abstract：:To maintain cellular homeostasis, the levels of transmembrane receptors found on the plasma membrane must be tightly regulated. Endocytosis of activated receptors and the eventual degradation of these transmembrane proteins in the lysosome serve a vital role in maintaining the plasma membrane receptor levels as well a...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)03049-1
更新日期：2005-01-01 00:00:00
abstract：:Intraflagellar transport (IFT) involves the movement of large proteinaceous particles or trains along the length of ciliary and flagellar axonemal microtubules. The particles contain multiple copies of two protein complexes. As isolated from the flagellated model organism, Chlamydomonas reinhardtii, IFT A contains 6 d...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-397945-2.00010-X
更新日期：2013-01-01 00:00:00
abstract：:Ubiquitin-like protein (Ubl) ligation is common to diverse archaea and targets many cellular pathways, including those associated with sulfur mobilization, and also tags proteins as substrates for degradation by the proteasome. Here we highlight protocols to assay proteasome function and Ubl ligation in archaea. A cha...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2018.12.036
更新日期：2019-01-01 00:00:00
abstract：:Autophagy is the major cellular process of degradation and is modulated by several signaling pathways. Phosphatidylinositol 3-kinase (PtdIns3K) class III (Vps34) and PtdIns3K class I regulate the autophagy pathway positively and negatively, respectively. Both classes of PtdIns3K participate in the synthesis of phospha...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2016.09.060
更新日期：2017-01-01 00:00:00
abstract：:Dynamin I is a large GTPase enzyme required in membrane constriction and fission during multiple forms of endocytosis. The first method described here is for the rapid purification of native dynamin from peripheral membrane extracts of sheep brain using ammonium sulfate precipitation and affinity purification on recom...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)04049-8
更新日期：2005-01-01 00:00:00
abstract：:Pulsed electron double resonance technique, also known as double electron-electron resonance, jointly with site-directed spin labeling (SDSL) have been used extensively for studying structures and structural change. During the last decades, significant enhancements have been made by optimization of the experimental pr...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2015.05.027
更新日期：2015-01-01 00:00:00
abstract：:Isotopic enrichment of biomacromolecules is a widely used technique that enables the investigation of the structural and dynamic properties to provide information not accessible with natural abundance isotopic composition. This study reports an approach for deuterium incorporation into bacterial cellulose. A media for...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2015.08.031
更新日期：2015-01-01 00:00:00
abstract：:As for other GPCRs, the oligomerization of glycoprotein hormone receptors (GPHRs) appears as critical event for receptor function. By means of modern techniques based on the BRET or FRET principle, GPHR oligomerization has been reported to explain several physiological and pathological conditions. In particular, the p...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-391862-8.00020-X
更新日期：2013-01-01 00:00:00
abstract：:Mass spectrometry (MS) has gradually replaced classical methods as a major tool in protein sequencing and characterization. However, the sample preparation repertoire has not changed very much; it has just been adjusted to the needs of the new analytical method. In this chapter frequently used in-solution enzymatic di...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)05003-2
更新日期：2005-01-01 00:00:00
abstract：:Precisely imaging and tracking dynamic biological processes in live cells are crucial for both fundamental research in life sciences and biomedical applications. Nonfluorescent nanoparticles are emerging as important optical probes in live-cell imaging because of their excellent photostability, large optical cross sec...
journal_title：Methods in enzymology
pub_type： 杂志文章,评审
doi：10.1016/B978-0-12-391857-4.00004-5
更新日期：2012-01-01 00:00:00
abstract：:Making error-free, custom DNA assemblies from potentially faulty building blocks is a fundamental challenge in synthetic biology. Here, we show how recursion can be used to address this challenge using a recursive procedure that constructs error-free DNA molecules and their libraries from error-prone synthetic oligonu...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-385120-8.00010-3
更新日期：2011-01-01 00:00:00
abstract：:Membrane proteins (MPs) are insoluble in aqueous media as a result of incompatibility between the hydrophilic property of the solvent molecules and the hydrophobic nature of MP surfaces, normally associated with lipid membranes. Amphipathic compounds are necessary for extraction of these macromolecules from the native...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2014.12.021
更新日期：2015-01-01 00:00:00
abstract：:Retinoic acid (RA), the biologically active metabolite of vitamin A, regulates a vast spectrum of biological processes, such as cell differentiation, proliferation, apoptosis, and morphogenesis. microRNAs (miRNAs) play a crucial role in regulating gene expression by binding to messenger RNA (mRNA) which leads to mRNA ...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2020.02.009
更新日期：2020-01-01 00:00:00
abstract：:Nonsense-mediated mRNA decay (NMD) is an evolutionarily conserved mechanism of specific degradation of transcripts with a premature stop codon. NMD eliminates aberrant mRNAs arising from mutations, alternative splicing, and other events in cells. In addition, many normal transcripts undergo NMD. Recent studies demonst...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2016.02.008
更新日期：2016-01-01 00:00:00
abstract：:Technical innovations in structural probing have drastically advanced the field of RNA structure analysis. These advances have led to parallel approaches developed in separate labs for analyzing RNA structure and dynamics. With the wealth of methodologies available, it can be difficult to determine which is best suite...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2019.04.002
更新日期：2019-01-01 00:00:00
abstract：:Cryptococcus neoformans is a yeastlike fungus that causes a lethal meningoencephalitis in a broad spectrum of immunocompromised patients and has become the most common cause of meningitis due to AIDS-related infections in Africa. Key to the development of new agents to control and prevent this infection is the identif...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(08)03222-9
更新日期：2008-01-01 00:00:00
abstract：:Autophagy is a major intracellular catabolic pathway induced in response to amino acid starvation. Recent findings implicate it in diverse physiological/pathophysiological events, such as protein and organelle turnover, development, aging, pathogen infection, cell death, and neurodegeneration. However, experimental me...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(08)03609-4
更新日期：2009-01-01 00:00:00
abstract：:Astrocytes are the most abundant cell type in the brain and are a crucial part of solving its mysteries. Originally assumed to be passive supporting cells, astrocyte's functions are now recognized to include active modulation and information processing at the neural synapse. The full extent of the astrocyte contributi...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2019.02.006
更新日期：2019-01-01 00:00:00
abstract：:Membrane-bound pyrophosphatases couple the hydrolysis of inorganic pyrophosphate to the pumping of ions (sodium or protons) across a membrane in order to generate an electrochemical gradient. This class of membrane protein is widely conserved across plants, fungi, archaea, and bacteria, but absent in multicellular ani...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2018.04.017
更新日期：2018-01-01 00:00:00
abstract：:In structural proteomics, it is necessary to efficiently screen in a high-throughput manner for the presence of stable structures in proteins that can be subjected to subsequent structure determination by X-ray or NMR spectroscopy. Here we illustrate that the (1)H chemical distribution in a protein as detected by (1)H...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)94006-8
更新日期：2005-01-01 00:00:00
abstract：:Research on circadian biology over the past decade has paid increasing attention to the photoreceptor mechanisms that align the molecular clock to the 24-h light/dark cycle, and some of the results to emerge are surprising. For example, the rods and cones within the mammalian eye are not required for entrainment. A po...
journal_title：Methods in enzymology
pub_type： 杂志文章,评审
doi：10.1016/S0076-6879(05)93037-1
更新日期：2005-01-01 00:00:00
abstract：:Specific recognition of mRNA 5' cap by eukaryotic initiation factor eIF4E is a rate-limiting step in the translation initiation. Structural determination of the eIF4E-cap complexes, as well as complexes of eIF4E with other proteins regulating its activity, requires complementary experiments that allow for energetic an...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(07)30009-8
更新日期：2007-01-01 00:00:00
abstract：:Dimethylsulfoniopropionate (DMSP) demethylase is a tetrahydrofolate-dependent enzyme that initiates the DMSP demethylation pathway in marine bacteria. This enzyme is important for understanding of organic sulfur flux from the oceans because it directs the sulfur from DMSP away from dimethylsulfide. This enzyme has bee...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2018.03.001
更新日期：2018-01-01 00:00:00
abstract：:In this chapter we have detailed a method that can be generalized to link virtually any DNA substrate to a chromatography matrix at its ends via an avidin-biotin linkage. We have used this technique to construct a left-handed Z-DNA column for the purpose of identification and purification of Z-DNA-binding proteins. Th...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/0076-6879(90)84290-w
更新日期：1990-01-01 00:00:00
abstract：:Benzylisoquinoline alkaloids (BIAs) constitute a diverse class of plant secondary metabolites that includes the opiate analgesics morphine and codeine. Collectively, BIAs exhibit a myriad of pharmacological activities, including antimicrobial, antitussive, antispasmodic, and anticancer properties. Despite 2500 known B...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2016.02.011
更新日期：2016-01-01 00:00:00
abstract：:Hematopoietic stem cells (HSCs) primarily reside in bone marrow, are defined by their ability to maintain blood homeostasis, and replenish themselves through self-renewal. Although HSC purification schemes vary from laboratory to laboratory, the resulting cell populations are similar, if not the same. This chapter wil...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(06)20011-9
更新日期：2006-01-01 00:00:00
abstract：:When yeast are faced with irreparable DNA damage, they will first arrest in G2/M, via the DNA damage checkpoint pathway, but will subsequently adapt to that arrest and resume division. Here, we summarize assays that we have used to examine checkpoint adaptation. Specifically, we discuss the merits of inducing DNA dama...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)09009-9
更新日期：2006-01-01 00:00:00
abstract：:The elucidation of structure-affinity relationships in EF-hand proteins requires a reliable assay of divalent ion affinity. In principle, isothermal titration calorimetry (ITC) should be capable of furnishing estimates for Ca2+- and Mg2+-binding constants in these systems. And because the method yields the binding ent...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(08)04210-9
更新日期：2009-01-01 00:00:00
