Whole-genome genotyping.

abstract：:We have developed two screening systems for isolating inhibitors that target bacterial two-component signal transduction: (1) a differential growth assay using a temperature-sensitive yycF mutant (CNM2000) of Bacillus subtilis, which is supersensitive to histidine kinase inhibitors, and (2) a high-throughput genetic s...

journal_title：Methods in enzymology

authors： Okada A,Gotoh Y,Watanabe T,Furuta E,Yamamoto K,Utsumi R

更新日期：2007-01-01 00:00:00

abstract：:Endocytosis and autophagy are key vesicular pathways involved in degradation and recycling of cellular material. Both degradative pathways finally fuse with lysosome but are indeed interconnected at several levels. In particular, the fusion between endosomes and autophagosomes can generate intermediate vesicles named ...

journal_title：Methods in enzymology

authors： Manil-Ségalen M,Culetto E,Legouis R,Lefebvre C

更新日期：2014-01-01 00:00:00

abstract：:Several genetically encoded fluorescent biosensors for Ras family GTPases have been developed that permit spatiotemporal analysis of the activation of these signaling molecules in living cells. We describe here the use of the simplest of these probes, the Ras binding domain (RBD) of selected effectors fused with green...

journal_title：Methods in enzymology

authors： Bivona TG,Quatela S,Philips MR

更新日期：2006-01-01 00:00:00

abstract：:Patient-derived xenografts are a useful tool in cancer immunology, as they allow researchers to study human cancers in vivo when starting with a relatively small amount of human tumor tissue. These models make it possible to study tumor cell-intrinsic changes that occur in response to external stimuli including cytoki...

journal_title：Methods in enzymology

authors： Cardenas JJ,Robles-Oteiza C,Politi K

更新日期：2020-01-01 00:00:00

abstract：:The number of genetically modified mouse lines has been increasing exponentially in the past few decades. In order to safeguard them from accidental loss and genetic drifting, to reduce animal housing cost, and to efficiently distribute them around the world, it is important to cryopreserve these valuable genetic reso...

journal_title：Methods in enzymology

authors： Takahashi H,Liu C

更新日期：2010-01-01 00:00:00

abstract：:Ecological samples of fungal associations pose particular challenges for nucleic acid extraction due to the presence of several genomes. Thorough examination of the samples prior to extraction is important to assess the risks of contamination. If manual separation of symbionts or their axenic cultivation is not feasib...

journal_title：Methods in enzymology

authors： Grube M

更新日期：2005-01-01 00:00:00

abstract：:Biominerals have complex and heterogeneous architectures, hence diffraction experiments with spatial resolutions between 500 nm and 10 μm are extremely useful to characterize them. X-ray beams in this size range are now routinely produced at many synchrotrons. This chapter provides a review of the different hard X-ray...

journal_title：Methods in enzymology

authors： Tamura N,Gilbert PU

更新日期：2013-01-01 00:00:00

abstract：:The study of model organisms in pure culture has provided detailed information about the physiology and biochemistry of nitrification and related processes. Metagenomic sequencing of environmental samples is providing information to what extent this understanding also applies to natural microbial communities. Here, we...

journal_title：Methods in enzymology

authors： Müller S,Strous M

更新日期：2011-01-01 00:00:00

abstract：:Guanine nucleotide dissociation inhibitor (GDI) is a central regulator of Rab GTPase family members. GDI recycles Rab proteins from the membrane and sequesters the inactive GDP-bound form of Rab in the cytosol for use in multiple rounds of transport. The balance between the membrane-bound form of Rab and the cytosolic...

journal_title：Methods in enzymology

authors： Chen CY,Sakisaka T,Balch WE

更新日期：2005-01-01 00:00:00

abstract：:Current methodologies available to quantify changes in mitochondrial turnover are limited to pulse-chase assays or specific assays that quantify mitophagy. Accordingly, new tools that can assess mitochondrial turnover are needed for the study of cellular, subcellular, and spatial parameters of mitochondrial turnover a...

journal_title：Methods in enzymology

authors： Trudeau KM,Gottlieb RA,Shirihai OS

更新日期：2014-01-01 00:00:00

abstract：:Formation of 5-methyluridine (ribothymidine) at position 54 of the T-psi loop of tRNA is catalyzed by site-specific tRNA methyltransferases (tRNA[uracil-54,C5]-MTases). In eukaryotes and many bacteria, the methyl donor for this reaction is generally S-adenosyl-L-methionine (S-AdoMet). However, in other bacteria, like ...

journal_title：Methods in enzymology

authors： Urbonavicius J,Brochier-Armanet C,Skouloubris S,Myllykallio H,Grosjean H

更新日期：2007-01-01 00:00:00

abstract：:Glycosylation is a complex form of protein modification occurring in the secretory pathway. The addition of N- and O-glycans affects intracellular processes like the folding and trafficking of most glycoproteins. To better understand the impact of glycosylation in protein folding and maturation, parameters like glycos...

journal_title：Methods in enzymology

authors： Hülsmeier AJ,Welti M,Hennet T

更新日期：2011-01-01 00:00:00

abstract：:The Gene Expression Omnibus (GEO) repository at the National Center for Biotechnology Information archives and freely distributes high-throughput molecular abundance data, predominantly gene expression data generated by DNA microarray technology. The database has a flexible design that can handle diverse styles of bot...

journal_title：Methods in enzymology

authors： Barrett T,Edgar R

更新日期：2006-01-01 00:00:00

abstract：:Complex RNA structures regulate many biological processes but are often too large for structure determination by nuclear magnetic resonance (NMR) methods. We determined the solution structure of domain II of the hepatitis C viral internal ribosome entry site (HCV IRES), a 25-kDa RNA, using a novel NMR approach. Conven...

journal_title：Methods in enzymology

authors： Lukavsky PJ,Puglisi JD

更新日期：2005-01-01 00:00:00

abstract：:Since its major launch into academia in the mid-1990s, spotted DNA microarray technology has expanded and matured into an important mainstream tool for genomic-scale gene expression studies across many species with many applications. Based on the principles of enzymatic nucleic acid labeling and DNA hybridization, the...

journal_title：Methods in enzymology

authors： Hager J

更新日期：2006-01-01 00:00:00

abstract：:The RNA-guided, sequence-specific endonuclease Cas9 has been widely adopted as genome engineering tool due to its efficiency and ease of use. Derived from the microbial CRISPR (clustered regularly interspaced short palindromic repeat) type II adaptive immune system, Cas9 has now been successfully engineered for genome...

journal_title：Methods in enzymology

authors： Trevino AE,Zhang F

更新日期：2014-01-01 00:00:00

abstract：:This chapter described the preparation and fractionation of libraries of M13 phage displaying proteins as fusions to the major coat protein. High titer (10(13) pfu/ml) phage libraries can readily be generated using a single vector and the level of display surpasses that of gene III fusion phage. Since the synthetic VI...

journal_title：Methods in enzymology

authors： Roberts BL,Markland W,Ladner RC

更新日期：1996-01-01 00:00:00

abstract：:PYP-phytochrome (Ppr) is a unique photoreceptor that contains a blue light-absorbing photoactive yellow protein (PYP) domain, a red light-absorbing phytochrome domain, and a histidine kinase domain. This chapter describes overexpression of Ppr in a strain of Escherichia coli that allows covalent attachment of substoic...

journal_title：Methods in enzymology

authors： Chung YH,Masuda S,Bauer CE

更新日期：2007-01-01 00:00:00

abstract：:The above methodology has been used to prepare a variety of PE-immunotoxins. These reagents are potent cytotoxic agents for cells in culture that bind the appropriate antibody and nontoxic when cells do not bind the antibody. PE-immunotoxins can be used to select for mutant cultured cells which lack the target of the ...

journal_title：Methods in enzymology

authors： FitzGerald DJ

更新日期：1987-01-01 00:00:00

abstract：:This protocol describes a general protein-to-protein cross-linking procedure using the water-soluble amine-reactive homobifunctional BS(3) (bis[sulfosuccinimidyl] suberate); however, the protocol can be easily adapted using other cross-linkers of similar properties. BS(3) is composed of two sulfo-NHS ester groups and ...

journal_title：Methods in enzymology

authors： Alegria-Schaffer A

更新日期：2014-01-01 00:00:00

abstract：:Desmoplakin (DP) is an obligate component of desmosomes, where it links the desmosomal cadherin/plakoglobin/plakophilin assembly to intermediate filaments. DP contains a large amino-terminal domain (DPNT) that binds to the cadherin/plakoglobin/plakophilin complex, a central coiled-coil domain that dimerizes the molecu...

journal_title：Methods in enzymology

authors： Choi HJ,Weis WI

更新日期：2016-01-01 00:00:00

abstract：:The circadian clock exists to synchronize inner physiology with the external world, allowing life to anticipate and adapt to the continual changes that occur in an organism's environment. The clock architecture is highly conserved, present in almost all major branches of life. Within eukaryotes, the filamentous fungus...

journal_title：Methods in enzymology

authors： Hurley J,Loros JJ,Dunlap JC

更新日期：2015-01-01 00:00:00

abstract：:Cell-free (CF) protein expression has emerged as one of the most efficient production platforms for membrane proteins. Central bottlenecks prevalent in conventional cell-based expression systems such as mistargeting, inclusion body formation, degradation as well as product toxicity can be addressed by taking advantage...

journal_title：Methods in enzymology

authors： Henrich E,Dötsch V,Bernhard F

更新日期：2015-01-01 00:00:00

abstract：:Nitric oxide formation in ischemia-reperfusion injury can be identified and measured directly using EPR of nitroso-heme complexes comprising NO bound to either Mb or Hb-Fe2+. This article described the successful use of this method to detect and quantify the generation of NO formed independently of nitric oxide syntha...

journal_title：Methods in enzymology

authors： Lepore DA

更新日期：2002-01-01 00:00:00

abstract：:This chapter presents methodologies for RNA extraction from soils coupled with competitive reverse transcription-polymerase chain reaction and capillary electrophoresis techniques. Combined, these approaches provide new capabilities to quantify gene expression in different environments and can aid our understanding of...

journal_title：Methods in enzymology

authors： Semrau JD,Han JI

更新日期：2005-01-01 00:00:00

abstract：:Two-dimensional polyacrylomide gel electrophoresis remains a popular and powerful tool for identifying proteins that are differentially expressed across treatment conditions. Due to the overwhelming number of proteins and the tremendous variation shown in gel images, the differential analysis of 2D gel images is chall...

journal_title：Methods in enzymology

authors： Li F,Seillier-Moiseiwitsch F

更新日期：2011-01-01 00:00:00

abstract：:Quantitative real-time polymerase chain reaction (qRT-PCR) is a flexible and scalable method for analyzing transcript abundance that can be used at a single gene or high-throughput (>100 genes) level. Information obtained from this technique can be used as an indicator of potential regulation of glycosylation at the t...

journal_title：Methods in enzymology

authors： Nairn AV,dela Rosa M,Moremen KW

更新日期：2010-01-01 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

authors： Alfonso C,del Castillo U,Martín I,Muga A,Rivas G

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

authors： Davies BA,Carney DS,Horazdovsky BF

更新日期：2005-01-01 00:00:00

abstract：:The analysis of a reaction pathway by stopped-flow and chemical quench-flow methods provides the only means of directly measuring the rates of reaction at the active site of an enzyme. The interpretation of data from each technique is similar and involves solution of the differential equations describing the approach ...

journal_title：Methods in enzymology

authors： Johnson KA

更新日期：1986-01-01 00:00:00