Generation of Mutant mASIC1a Constructs

The complementary DNA (cDNA) encoding mouse ASIC1a (mASIC1a) was used as previously described (Lynagh et al., 2017 (link)). Site-directed mutagenesis was performed using PfuUltraII Fusion polymerase (Agilent) and custom DNA mutagenesis primers (Eurofins Genomics). All sequences were confirmed by sequencing of the full coding frame (Eurofins Genomics or Macrogen). cDNAs were linearized with EcoRI, and capped cRNA was transcribed with the Ambion mMESSAGE mMACHINE SP6 kit (Thermo Fisher Scientific).

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Heusser S.A., Borg C.B., Colding J.M, & Pless S.A. (2022). Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition. eLife, 11, e73384.

Publication 2022

PfuUltraII Fusion polymerase custom DNA mutagenesis primers Ambion mMESSAGE mMACHINE SP6 kit

Cdnas Crna Dna primers Ecori Frame Mouse Mutagenesis Site directed mutagenesis

Corresponding Organization :

Other organizations : University of Copenhagen

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Variable analysis

independent variables

Site-directed mutagenesis

dependent variables

Sequences of the full coding frame

control variables

PfuUltraII Fusion polymerase
Custom DNA mutagenesis primers
EcoRI linearization
Ambion mMESSAGE mMACHINE SP6 kit

controls

Positive control: None mentioned
Negative control: None mentioned

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