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Methods Mol Biol
2022 Jan 01;2510:157-192. doi: 10.1007/978-1-0716-2384-8_9.
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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
BERTANI,
Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli.
1951,
Pubmed
Becker,
The P2X7 carboxyl tail is a regulatory module of P2X7 receptor channel activity.
2008,
Pubmed
,
Xenbase
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,
Substitution of chloroform by bromo-chloropropane in the single-step method of RNA isolation.
1995,
Pubmed
Chomczynski,
The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction: twenty-something years on.
2006,
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
