Abstract:
:Photoconvertible fluorescent proteins, such as Kaede, can be switched irreversibly from their native color to a new one. This property can be exploited to visualize de novo mRNA translation, because newly synthesized proteins can be distinguished from preexisting ones by their color. In this protocol, Kaede cDNA linked to the 3' untranslated region (UTR) of beta-actin is delivered into cells fated to become the retina by injection into Xenopus blastomeres. Brief exposure (6-10 s) to UV light (350-410 nm) of Kaede-positive retinal axons/growth cones efficiently converts Kaede from its native green fluorescence to red. The reappearance of the green signal reports the synthesis of new Kaede protein. This approach can be used to investigate the spatiotemporal control of translation of specific mRNAs in response to external stimuli and to test the efficiency of full-length versus mutant UTRs. The 3-d protocol can be adapted for broad use with other photoactivatable fluorescent proteins.
journal_name
Nat Protocjournal_title
Nature protocolsauthors
Leung KM,Holt CEdoi
10.1038/nprot.2008.113subject
Has Abstractpub_date
2008-01-01 00:00:00pages
1318-27issue
8eissn
1754-2189issn
1750-2799pii
nprot.2008.113journal_volume
3pub_type
杂志文章相关文献
Nature Protocols文献大全abstract::Hypochlorous acid (HOCl) is a critical member of the reactive oxygen species (ROS) produced by immune cells to fight infections. On the other hand, HOCl in homeostasis causes oxidative damage to biomolecules and is linked to many diseases, including inflammatory, neurodegenerative, and cardiovascular diseases. Herein,...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/s41596-018-0041-6
更新日期:2018-10-01 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
pub_type: 杂志文章
doi:10.1038/nprot.2006.186
更新日期:2006-01-01 00:00:00
abstract::The research community is in a race to understand the molecular mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, to repurpose currently available antiviral drugs and to develop new therapies and vaccines against coronavirus disease 2019 (COVID-19). One major challenge in achieving ...
journal_title:Nature protocols
pub_type: 杂志文章,评审
doi:10.1038/s41596-020-00403-2
更新日期:2020-12-01 00:00:00
abstract::Ex vivo perfusion systems offer a reliable, reproducible method for studying acute physiological responses of an organ to various environmental manipulations. Unlike in vitro culture systems, the cellular organization, compartmentalization and three-dimensional structure of ex vivo-perfused organs are maintained. Thes...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2012.144
更新日期:2013-01-01 00:00:00
abstract::Social recognition (SR) enables rodents to distinguish between familiar and novel conspecifics, largely through individual odor cues. SR tasks utilize the tendency for a male to sniff and interact with a novel individual more than a familiar individual. Many paradigms have been used to study the roles of the neuropept...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2009.141
更新日期:2009-01-01 00:00:00
abstract::Accurate analysis of DNA sequence variation in not only humans and animals but also other organisms has played a significant role in expanding our knowledge about genetic variety and diversity in a number of different biological areas. The search for an understanding of the causes of genetic variants and mutations has...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2007.407
更新日期:2007-01-01 00:00:00
abstract::The elucidation of protein-protein interaction networks is a crucial task in the postgenomic era. In this protocol, we describe our approach to discover protein-protein interactions using the surface plasmon resonance technique coupled to mass spectrometry (MS). A peptide or a protein is immobilized on a sensor chip a...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2009.84
更新日期:2009-01-01 00:00:00
abstract::A big challenge in proteomics is the identification of cell-type-specific proteomes in vivo. This protocol describes how to label, purify and identify cell-type-specific proteomes in living mice. To make this possible, we created a Cre-recombinase-inducible mouse line expressing a mutant methionyl-tRNA synthetase (L27...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/s41596-018-0106-6
更新日期:2019-02-01 00:00:00
abstract::Cyclodepsipeptides are cyclic peptides in which at least one amide link on the backbone is replaced with an ester link. These natural products present a high structural diversity that corresponds to a broad range of biological activities. Therefore, they are very promising pharmaceutical candidates. Most of the cyclod...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2016.116
更新日期:2016-10-01 00:00:00
abstract::The unprecedented increase in the throughput of DNA sequencing driven by next-generation technologies now allows efficient analysis of the complete protein-coding regions of genomes (exomes) for multiple samples in a single sequencing run. However, sample preparation and targeted enrichment of multiple samples has bec...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2011.396
更新日期:2011-11-03 00:00:00
abstract::AMPylation (adenylylation) has been recognized as an important post-translational modification that is used by pathogens to regulate host cellular proteins and their associated signaling pathways. AMPylation has potential functions in various cellular processes, and it is widely conserved across both prokaryotes and e...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2015.044
更新日期:2015-05-01 00:00:00
abstract::The telomeric repeat amplification protocol (TRAP) is a two-step process for analyzing telomerase activity in cell or tissue extracts. Recent modifications of this sensitive assay include elimination of radioactivity by using a fluorescently labeled primer instead of a radiolabeled primer. In addition, the TRAP assay ...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2006.239
更新日期:2006-01-01 00:00:00
abstract::Carbohydrate microarrays have received considerable attention as an advanced technology for the rapid analysis of carbohydrate-protein interactions. This protocol provides detailed procedures for the preparation of carbohydrate microarrays by immobilizing hydrazide-conjugated carbohydrates on epoxide-derivatized glass...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2007.373
更新日期:2007-01-01 00:00:00
abstract::Forebrain γ-aminobutyric acid (GABA) interneurons have crucial roles in high-order brain function via modulating network activities and plasticity, and they are implicated in many psychiatric disorders. Availability of enriched functional human forebrain GABA interneurons, especially those from people affected by GABA...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2013.106
更新日期:2013-09-01 00:00:00
abstract::This protocol represents a novel enzyme-luminescence method to detect dopamine sensitively and rapidly with high temporal resolution. In principle, dopamine is first oxidized with tyramine oxidase to produce H(2)O(2), and then the produced H(2)O(2) reacts with luminol to generate chemiluminescence in the presence of h...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2008.158
更新日期:2008-01-01 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
pub_type: 杂志文章
doi:10.1038/nprot.2007.307
更新日期:2007-01-01 00:00:00
abstract::Methods for installing natural and unnatural amino acids and their modifications into proteins in a benign and precise manner are highly sought-after in protein science. Here we describe a protocol for 'post-translational mutagenesis' that enables the programmed installation of protein side chains through the use of r...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2017.087
更新日期:2017-10-01 00:00:00
abstract::The synthesis of an azobenzene amino acid (aa) for use as a photo-inducible conformational switch in polypeptides is described. The compound can be easily incorporated into an aa sequence by solid-phase peptide synthesis using standard 9-fluorenylmethoxycarbonyl methods. A reversible conformational change of the pepti...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2006.488
更新日期:2007-01-01 00:00:00
abstract::The plant cell wall is a chemically complex structure composed mostly of polysaccharides. Detailed analyses of these cell wall polysaccharides are essential for our understanding of plant development and for our use of plant biomass (largely wall material) in the food, agriculture, fabric, timber, biofuel and biocompo...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2012.081
更新日期:2012-09-01 00:00:00
abstract::This protocol describes a simple silver staining method used to visualize DNA fragments and other organic molecules with unsurpassed detail following traditional polyacrylamide gel electrophoresis (PAGE). Sensitivity rivals radioisotopic methods and DNA in the picogram range can be reliably detected. The described pro...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2007.330
更新日期:2007-01-01 00:00:00
abstract::The Golgi apparatus undergoes extensive disassembly during mitosis and reassembly in post-mitotic daughter cells. This process has been mimicked in vitro by treating Golgi membranes with mitotic and interphase cytosol. To determine the minimal machinery that controls the morphological change, we have developed a defin...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2010.38
更新日期:2010-04-01 00:00:00
abstract::This protocol describes a simple but robust microfluidic assay combining three-dimensional (3D) and two-dimensional (2D) cell culture. The microfluidic platform comprises hydrogel-incorporating chambers between surface-accessible microchannels. By using this platform, well-defined biochemical and biophysical stimuli c...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2012.051
更新日期:2012-06-07 00:00:00
abstract::The pig has emerged as an important large animal model in biomedical and pharmaceutical research. We describe a protocol for high-efficiency germline transgenesis and sustained transgene expression in pigs by using the Sleeping Beauty (SB) transposon system. The protocol is based on co-injection of a plasmid encoding ...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2014.010
更新日期:2014-04-01 00:00:00
abstract::Freestanding plasmonic nanoparticle (NP) superlattice sheets are novel 2D nanomaterials with tailorable properties that enable their use for broad applications in sensing, anticounterfeit measures, ionic gating, nanophotonics and flat lenses. We recently developed a robust, yet general, two-step drying-mediated approa...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/s41596-019-0200-4
更新日期:2019-09-01 00:00:00
abstract::The systematic mapping of protein interactions by bait-prey techniques, including affinity purification-mass spectrometry or the yeast two-hybrid system, contributes a unique and relevant perspective on the comprehensive picture of cellular machines. We describe here a protocol for statistical analysis of node-and-edg...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2009.26
更新日期:2009-01-01 00:00:00
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
pub_type: 杂志文章
doi:10.1038/nprot.2014.105
更新日期:2014-07-01 00:00:00
abstract::The Buccal Micronucleus Cytome (BMCyt) assay is a minimally invasive method for studying DNA damage, chromosomal instability, cell death and the regenerative potential of human buccal mucosal tissue. This method is increasingly used in molecular epidemiological studies for investigating the impact of nutrition, lifest...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2009.53
更新日期:2009-01-01 00:00:00
abstract::We describe the implementation and use of an adaptive imaging framework for optimizing spatial resolution and signal strength in a light-sheet microscope. The framework, termed AutoPilot, comprises hardware and software modules for automatically measuring and compensating for mismatches between light-sheet and detecti...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/s41596-018-0043-4
更新日期:2018-11-01 00:00:00
abstract::Positron emission tomography (PET) is a sensitive and noninvasive imaging method that is widely used to explore molecular events in living subjects. PET can precisely and quantitatively evaluate cellular apoptosis, which has a crucial role in various physiological and pathological processes. In this protocol, we descr...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2015.048
更新日期:2015-05-01 00:00:00
abstract::This protocol describes the efficient, generally applicable Ullmann coupling reaction of bromaminic acid with alkyl- or aryl-amines in phosphate buffer under microwave irradiation using elemental copper as a catalyst. The reaction leads to a number of biologically active compounds. As a prototypical example, the synth...
journal_title:Nature protocols
pub_type: 杂志文章
doi:10.1038/nprot.2010.63
更新日期:2010-05-01 00:00:00