Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Methods Mol Biol
2022 Jan 01;2510:157-192. doi: 10.1007/978-1-0716-2384-8_9.
Show Gene links
Show Anatomy links
Established Protocols for cRNA Expression and Voltage-Clamp Characterization of the P2X7 Receptor in Xenopus laevis Oocytes.
Schmalzing G, Markwardt F.
???displayArticle.abstract??? P2X7 receptors (P2X7Rs) are fast ATP4--gated ion channels that, like other members of the P2X receptor family, function as homotrimers. A high-resolution cryo-EM structure of the full-length rat P2X7R is available. Using voltage-clamp experiments in Xenopus laevis oocytes, even the earliest steps of P2X7R activation can be quantitatively recorded in the millisecond range. Site-directed mutagenesis combined with voltage-clamp recordings can reveal residues and domains of the P2X7R involved in ATP4- binding, gating (i.e., opening and closing of the channel pore) and ion selectivity. We present here proven voltage-clamp protocols that take into account requirements that are important at the levels of cDNA and vector sequences, cRNA synthesis, and Xenopus laevis oocyte isolation for reliable results.
BARLOW,
A SIMPLE METHOD FOR THE QUANTITATIVE ISOLATION OF UNDEGRADED HIGH MOLECULAR WEIGHT RIBONUCLEIC ACID.
1963, Pubmed
BARLOW,
A SIMPLE METHOD FOR THE QUANTITATIVE ISOLATION OF UNDEGRADED HIGH MOLECULAR WEIGHT RIBONUCLEIC ACID.
1963,
Pubmed Becker,
The P2X7 carboxyl tail is a regulatory module of P2X7 receptor channel activity.
2008,
Pubmed
,
Xenbase BERTANI,
Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli.
1951,
Pubmed Brawerman,
A procedure for the isolation of mammalian messenger ribonucleic acid.
1972,
Pubmed Bretschneider,
Drug-dependent ion channel gating by application of concentration jumps using U-tube technique.
1999,
Pubmed Chomczynski,
The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction: twenty-something years on.
2006,
Pubmed Chomczynski,
Substitution of chloroform by bromo-chloropropane in the single-step method of RNA isolation.
1995,
Pubmed Cunningham,
Use of inorganic pyrophosphatase to improve the yield of in vitro transcription reactions catalyzed by T7 RNA polymerase.
1990,
Pubmed Dascal,
The use of Xenopus oocytes for the study of ion channels.
1987,
Pubmed
,
Xenbase De Robertis,
Coupled transcription-translation of DNA injected into Xenopus oocytes.
1977,
Pubmed
,
Xenbase Di Virgilio,
The Elusive P2X7 Macropore.
2018,
Pubmed Flittiger,
Effects of protons on macroscopic and single-channel currents mediated by the human P2X7 receptor.
2010,
Pubmed
,
Xenbase Green,
Isolation of High-Molecular-Weight DNA Using Organic Solvents.
2017,
Pubmed Grudzien-Nogalska,
Synthesis of anti-reverse cap analogs (ARCAs) and their applications in mRNA translation and stability.
2007,
Pubmed Hamill,
Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches.
1981,
Pubmed Hattori,
Molecular mechanism of ATP binding and ion channel activation in P2X receptors.
2012,
Pubmed Hausmann,
ATP binding site mutagenesis reveals different subunit stoichiometry of functional P2X2/3 and P2X2/6 receptors.
2012,
Pubmed
,
Xenbase Henderson,
Cap 1 Messenger RNA Synthesis with Co-transcriptional CleanCap® Analog by In Vitro Transcription.
2021,
Pubmed Hernández,
Conservation and Variability of the AUG Initiation Codon Context in Eukaryotes.
2019,
Pubmed Illes,
Update of P2X receptor properties and their pharmacology: IUPHAR Review 30.
2021,
Pubmed Kearse,
Non-AUG translation: a new start for protein synthesis in eukaryotes.
2017,
Pubmed Kottur,
Pyrophosphate hydrolysis is an intrinsic and critical step of the DNA synthesis reaction.
2018,
Pubmed Kowalski,
Flexible subunit stoichiometry of functional human P2X2/3 heteromeric receptors.
2015,
Pubmed
,
Xenbase Kozak,
Adherence to the first-AUG rule when a second AUG codon follows closely upon the first.
1995,
Pubmed Krieg,
Functional messenger RNAs are produced by SP6 in vitro transcription of cloned cDNAs.
1984,
Pubmed
,
Xenbase Kubick,
The effect of anions on the human P2X7 receptor.
2011,
Pubmed
,
Xenbase Leppek,
Functional 5' UTR mRNA structures in eukaryotic translation regulation and how to find them.
2018,
Pubmed McCarthy,
Full-Length P2X7 Structures Reveal How Palmitoylation Prevents Channel Desensitization.
2019,
Pubmed
,
Xenbase McCaughan,
Translational termination efficiency in mammals is influenced by the base following the stop codon.
1995,
Pubmed Melton,
Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter.
1984,
Pubmed Mertz,
Purified DNAs are transcribed after microinjection into Xenopus oocytes.
1977,
Pubmed
,
Xenbase Munroe,
Tales of poly(A): a review.
1990,
Pubmed Nicke,
Homotrimeric complexes are the dominant assembly state of native P2X7 subunits.
2008,
Pubmed
,
Xenbase Park,
mRNA vaccines for COVID-19: what, why and how.
2021,
Pubmed Pasquinelli,
Reverse 5' caps in RNAs made in vitro by phage RNA polymerases.
1995,
Pubmed
,
Xenbase Pippel,
Localization of the gate and selectivity filter of the full-length P2X7 receptor.
2017,
Pubmed Riedel,
Kinetics of P2X7 receptor-operated single channels currents.
2007,
Pubmed
,
Xenbase Schenborn,
A novel transcription property of SP6 and T7 RNA polymerases: dependence on template structure.
1985,
Pubmed Schneider,
Interaction of Purinergic P2X4 and P2X7 Receptor Subunits.
2017,
Pubmed
,
Xenbase Schubert,
Multiple ligand-ion solutions: a guide for solution preparation and computer program understanding.
1996,
Pubmed Shatkin,
Capping of eucaryotic mRNAs.
1976,
Pubmed Simonetti,
Structural Insights into the Mammalian Late-Stage Initiation Complexes.
2020,
Pubmed Stepinski,
Synthesis and properties of mRNAs containing the novel "anti-reverse" cap analogs 7-methyl(3'-O-methyl)GpppG and 7-methyl (3'-deoxy)GpppG.
2001,
Pubmed Stolz,
Homodimeric anoctamin-1, but not homodimeric anoctamin-6, is activated by calcium increases mediated by the P2Y1 and P2X7 receptors.
2015,
Pubmed
,
Xenbase Tan,
Salt contribution to RNA tertiary structure folding stability.
2011,
Pubmed Thakur,
eIF2α interactions with mRNA control accurate start codon selection by the translation preinitiation complex.
2020,
Pubmed Walker,
RNA purification--precipitation methods.
2013,
Pubmed
