Expander: from expression microarrays to networks and functions.

abstract：:Overexpression screens can be used to explore gene function in Drosophila melanogaster, but to demonstrate their full potential, comprehensive and systematic collections of fly strains are required. Here we provide a protocol for high-throughput cloning of Drosophila open-reading frames (ORFs) that are regulated by up...

journal_title：Nature protocols

authors： Bischof J,Sheils EM,Björklund M,Basler K

更新日期：2014-07-01 00:00:00

abstract：:High-throughput transcriptional analysis has unveiled a myriad of novel RNAs. However, technical constraints in RNA sequencing library preparation and platform performance hamper the identification of rare transcripts contained within the RNA repertoire. Herein we present targeted-RNA directional sequencing (TARDIS), ...

journal_title：Nature protocols

authors： Portal MM,Pavet V,Erb C,Gronemeyer H

更新日期：2015-12-01 00:00:00

abstract：:MicroRNAs (miRNAs) are short, about 21 nucleotides in length, noncoding, regulatory RNA molecules representing a new layer in post-transcriptional regulation of gene expression. Intensive miRNA research has necessitated the development of effective miRNA detection methods such as northern analyses, quantitative real-t...

journal_title：Nature protocols

authors： Várallyay E,Burgyán J,Havelda Z

更新日期：2008-01-01 00:00:00

abstract：:Here we present a protocol for the extraction of RNA from Dictyostelium discoideum. Dictyostelium is a social amoeba that undergoes a basic developmental program, and therefore analysis of RNA levels over a time course is a commonly used technique. This procedure is similar to other guanidine thiocyanate-based methods...

journal_title：Nature protocols

authors： Pilcher KE,Gaudet P,Fey P,Kowal AS,Chisholm RL

更新日期：2007-01-01 00:00:00

abstract：:Methods development in chromatography is a time-consuming, trial-and-error process that requires laborious experimentation. We describe a high-throughput screening (HTS) protocol for the rapid identification of chromatographic steps for protein purification from cell-free expression broths. Broths containing the prote...

journal_title：Nature protocols

authors： Rege K,Heng M

更新日期：2010-03-01 00:00:00

abstract：:In this report, we describe a two-step protocol for labeling of an affinity-purified antibody to biotin with horseradish peroxidase (HRP) using cyanuric chloride (CC) as a bridge. The enzyme was first modified with CC, and following chromatography on a PD-10 column, the activated HRP was incubated with the antibody to...

journal_title：Nature protocols

authors： Abuknesha RA,Jeganathan F,Wu J,Baalawy Z

更新日期：2009-01-01 00:00:00

abstract：:The point mutational spectrum over nearly any 75- to 250-bp DNA sequence isolated from cells, tissues or large populations may be discovered using denaturing capillary electrophoresis (DCE). A modification of the standard DCE method that uses cycling temperature (e.g., +/-5 degrees C), CyDCE, permits optimal resolutio...

journal_title：Nature protocols

authors： Ekstrøm PO,Khrapko K,Li-Sucholeiki XC,Hunter IW,Thilly WG

更新日期：2008-01-01 00:00:00

abstract：:The new generation of synchrotrons and microfocused beamlines has enabled great progress in X-ray protein crystallography, resulting in new 3D atomic structures for proteins of high interest to the pharmaceutical industry and life sciences. It is, however, often still challenging to produce protein crystals of suffici...

journal_title：Nature protocols

authors： Pechkova E,Nicolini C

更新日期：2017-12-01 00:00:00

abstract：:Prediction of protein-protein interactions at the structural level on the proteome scale is important because it allows prediction of protein function, helps drug discovery and takes steps toward genome-wide structural systems biology. We provide a protocol (termed PRISM, protein interactions by structural matching) f...

journal_title：Nature protocols

authors： Tuncbag N,Gursoy A,Nussinov R,Keskin O

更新日期：2011-08-11 00:00:00

abstract：:Protein ubiquitination is a versatile protein modification that regulates virtually all cellular processes. This versatility originates from polyubiquitin chains, which can be linked in eight distinct ways. The combinatorial complexity of eight linkage types in homotypic (one chain type per polymer) and heterotypic (m...

journal_title：Nature protocols

authors： Hospenthal MK,Mevissen TET,Komander D

更新日期：2015-02-01 00:00:00

abstract：:The CRISPR-Cas9 genome-editing tool and the availability of whole-genome sequences from plant species have revolutionized our ability to introduce targeted mutations into important crop plants, both to explore genetic changes and to introduce new functionalities. Here, we describe protocols adapting the CRISPR-Cas9 sy...

journal_title：Nature protocols

authors： Osakabe Y,Liang Z,Ren C,Nishitani C,Osakabe K,Wada M,Komori S,Malnoy M,Velasco R,Poli M,Jung MH,Koo OJ,Viola R,Nagamangala Kanchiswamy C

更新日期：2018-12-01 00:00:00

abstract：:High-throughput sequencing of the variable domains of immune receptors (antibodies and T cell receptors (TCRs)) is of key importance in the understanding of adaptive immune responses in health and disease. However, the sequencing of both immune receptor chains (VH+VL or TCRβ/δ+TCRα/γ) at the single-cell level for typi...

journal_title：Nature protocols

authors： McDaniel JR,DeKosky BJ,Tanno H,Ellington AD,Georgiou G

更新日期：2016-03-01 00:00:00

abstract：:We describe procedures for the synthesis of a fluorescent pyrimidine analog and its site-specific incorporation into a DNA oligomer. The 5'-protected and 3'-activated nucleoside 4 is synthesized in three steps with an overall yield of 40%. Site-specific incorporation into a DNA oligomer occurs with greater than 88% co...

journal_title：Nature protocols

authors： Greco NJ,Tor Y

更新日期：2007-01-01 00:00:00

abstract：:Human embryonic stem (hES) cells are self-renewing, pluripotent cells that are valuable research tools and hold promise for use in regenerative medicine. Most hES cell lines are derived from cryopreserved human embryos that were created during in vitro fertilization (IVF) and are in excess of clinical need. Embryos th...

journal_title：Nature protocols

authors： Lerou PH,Yabuuchi A,Huo H,Miller JD,Boyer LF,Schlaeger TM,Daley GQ

更新日期：2008-01-01 00:00:00

abstract：:The human stomach contains two primary domains: the corpus, which contains the fundic epithelium, and the antrum. Each of these domains has distinct cell types and functions, and therefore each presents with unique disease pathologies. Here, we detail two protocols to differentiate human pluripotent stem cells (hPSCs)...

journal_title：Nature protocols

authors： Broda TR,McCracken KW,Wells JM

更新日期：2019-01-01 00:00:00

abstract：:A fluorescence-based microarray technique that does not require target DNA labeling is detailed. This 'label-free' approach utilizes a cationic, water-soluble conjugated polymer PFBT (poly[9,9'-bis(6''-(N,N,N-trimethylammonium)hexyl)fluorene-co-alt-4,7-(2,1,3-benzothiadiazole) dibromide]), and neutral PNA (peptide nuc...

journal_title：Nature protocols

authors： Sun C,Gaylord BS,Hong JW,Liu B,Bazan GC

更新日期：2007-01-01 00:00:00

abstract：:Cap-analysis gene expression (CAGE) provides accurate high-throughput measurement of RNA expression. CAGE allows mapping of all the initiation sites of both capped coding and noncoding RNAs. In addition, transcriptional start sites within promoters are characterized at single-nucleotide resolution. The latter allows t...

journal_title：Nature protocols

authors： Takahashi H,Lassmann T,Murata M,Carninci P

更新日期：2012-02-23 00:00:00

abstract：:RNase H-dependent PCR-enabled T-cell receptor sequencing (rhTCRseq) can be used to determine paired alpha/beta T-cell receptor (TCR) clonotypes in single cells or perform alpha and beta TCR repertoire analysis in bulk RNA samples. With the enhanced specificity of RNase H-dependent PCR (rhPCR), it achieves TCR-specific...

journal_title：Nature protocols

authors： Li S,Sun J,Allesøe R,Datta K,Bao Y,Oliveira G,Forman J,Jin R,Olsen LR,Keskin DB,Shukla SA,Wu CJ,Livak KJ

更新日期：2019-08-01 00:00:00

abstract：:The transmembrane (TM) anchors of cell surface proteins have been one of the 'blind spots' in structural biology because they are generally very hydrophobic, sometimes dynamic, and thus difficult targets for structural characterization. A plethora of examples show these membrane anchors are not merely anchors but can ...

journal_title：Nature protocols

authors： Fu Q,Piai A,Chen W,Xia K,Chou JJ

更新日期：2019-08-01 00:00:00

abstract：:This protocol describes an optimized method for direct in vitro monitoring of homo- and heterotypic axon-axon interactions involved in the developmental assembly of neural circuits. The assay exploits a classical example of heterotypic axonal interactions by modeling the sequential extension of spinal motor and somato...

journal_title：Nature protocols

authors： Wang L,Marquardt T

更新日期：2012-01-26 00:00:00

abstract：:The small ubiquitin-like modifiers (SUMOs) are posttranslationally conjugated to eukaryotic cellular proteins with generally unpredictable consequences. SUMO substrates are found in many cellular systems, and functional analysis has revealed that substrate SUMOylation often has an important role in their regulation. H...

journal_title：Nature protocols

authors： Tatham MH,Rodriguez MS,Xirodimas DP,Hay RT

更新日期：2009-01-01 00:00:00

abstract：:The analysis of large microbiome data sets holds great promise for the delineation of the biological and metabolic functioning of living organisms and their role in the environment. In the midst of this genomic puzzle, viruses, especially those that infect microbial communities, represent a major reservoir of genetic ...

journal_title：Nature protocols

authors： Paez-Espino D,Pavlopoulos GA,Ivanova NN,Kyrpides NC

更新日期：2017-08-01 00:00:00

abstract：:Single-cell electroporation allows transfection of plasmid DNA or macromolecules into individual living cells using modified patch electrodes and common electrophysiological equipment. This protocol is optimized for rapid in vivo electroporation of Xenopus laevis tadpole brains with DNA, dextrans, morpholinos and comb...

journal_title：Nature protocols

authors： Bestman JE,Ewald RC,Chiu SL,Cline HT

更新日期：2006-01-01 00:00:00

abstract：:This protocol measures externalization of aminophospholipids (APLs) to the outside of the plasma membrane using mass spectrometry (MS). APL externalization occurs in numerous events, and it is relevant for transplant medicine, immunity and cancer. In this protocol, externalized APLs are chemically modified by using a ...

journal_title：Nature protocols

authors： Thomas CP,Clark SR,Hammond VJ,Aldrovandi M,Collins PW,O'Donnell VB

更新日期：2014-01-01 00:00:00

abstract：:Blood circulating endothelial cells (CECs) and circulating hematopoietic progenitor cells (CPCs) represent two cell populations that are thought to play important roles in tissue vascularization. CECs and CPCs are currently studied as surrogate markers in patients for more than a dozen pathologies, including heart dis...

journal_title：Nature protocols

authors： Duda DG,Cohen KS,Scadden DT,Jain RK

更新日期：2007-01-01 00:00:00

abstract：:The protein called 'small ubiquitin-like modifier' (SUMO) is post-translationally linked to target proteins at the ɛ-amino group of lysine residues. This 'SUMOylation' alters the behavior of the target protein, a change that is utilized to regulate diverse cellular processes. Understanding the target-specific conseque...

journal_title：Nature protocols

authors： Tammsalu T,Matic I,Jaffray EG,Ibrahim AF,Tatham MH,Hay RT

更新日期：2015-09-01 00:00:00

abstract：:Improvement of cellular uptake and cellular localization is still one of the main obstacles to the development of antisense-antigene therapeutics, including peptide nucleic acid (PNA). Cell-penetrating peptides (CPPs) such as Tat peptide and polyarginine have been widely used to improve the cellular uptake of PNA and ...

journal_title：Nature protocols

authors： Shiraishi T,Nielsen PE

更新日期：2006-01-01 00:00:00

abstract：:A procedure for the synthesis of N-succinimidyl 3-iodobenzoate labeled with any iodine isotope ([*I]SIB), which is an agent used in the radioiodination of proteins and peptides, from its tin precursor N-succinimidyl 3-(tri-n-butylstannyl)benzoate (STB) is described. Also included are protocols for the synthesis of an ...

journal_title：Nature protocols

authors： Vaidyanathan G,Zalutsky MR

更新日期：2006-01-01 00:00:00

abstract：:Targeted nucleases, including zinc-finger nucleases (ZFNs), transcription activator-like (TAL) effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9), have provided researchers with the ability to manipulate nearly any genomic sequence in hu...

journal_title：Nature protocols

authors： Liu J,Gaj T,Yang Y,Wang N,Shui S,Kim S,Kanchiswamy CN,Kim JS,Barbas CF 3rd

更新日期：2015-11-01 00:00:00

abstract：:Influenza A viruses (IAVs) cause epidemics and pandemics that result in considerable financial burden and loss of human life. To manage annual IAV epidemics and prepare for future pandemics, an improved understanding of how IAVs emerge, transmit, cause disease and acquire pandemic potential is urgently needed. Fundame...

journal_title：Nature protocols

authors： Eisfeld AJ,Neumann G,Kawaoka Y

更新日期：2014-11-01 00:00:00