Merck

Protocol for Patch-Seq of Small Interneurons.

STAR protocols (2020-12-31)
Marcela Lipovsek, Lorcan Browne, Matthew S Grubb
ABSTRACT

Obtaining electrophysiological recordings and gene expression information from the same neuron (Patch-seq) brings forward a unique opportunity to study the transcriptional correlates of functional properties and vice versa. Here, we provide a detailed Patch-seq protocol tailored to the specialized demands of studying small interneurons. Focusing on the technically demanding process of transitioning between patch recordings and cell extraction, our protocol describes and troubleshoots steps for successfully collecting small interneurons, allowing for multi-modal Patch-seq interrogation of this crucial cell type.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Sodium bicarbonate, powder, BioReagent, for molecular biology, suitable for cell culture, suitable for insect cell culture
Sigma-Aldrich
DL-2-Amino-5-phosphonopentanoic acid, solid
Sigma-Aldrich
Magnesium chloride solution, for molecular biology, 1.00 M±0.01 M
Sigma-Aldrich
Adenosine 5′-triphosphate disodium salt solution, Crystalline ATP, HPLC purified, aqueous solution for RNA transcription
Sigma-Aldrich
Hydrochloric acid solution, 1.0 N, BioReagent, suitable for cell culture
Sigma-Aldrich
D-(+)-Glucose, BioXtra, ≥99.5% (GC)
Sigma-Aldrich
Acetic acid, glacial, ACS reagent, ≥99.7%
Sigma-Aldrich
Potassium chloride, ACS reagent, 99.0-100.5%
Sigma-Aldrich
Diethyl pyrocarbonate, 96% (NT)
Sigma-Aldrich
Potassium D-gluconate, ≥99%
Sigma-Aldrich
Sucrose, ≥99.5% (GC)
Sigma-Aldrich
Sodium phosphate monobasic, ReagentPlus®, ≥99.0%
Sigma-Aldrich
NBQX disodium salt hydrate, ≥98% (HPLC)
Sigma-Aldrich
Calcium chloride solution, BioUltra, for molecular biology, ~1 M in H2O
Sigma-Aldrich
Sucrose, BioUltra, for molecular biology, ≥99.5% (HPLC)
Sigma-Aldrich
SR-95531, ≥98% (HPLC), powder
Sigma-Aldrich
Sodium bicarbonate solution, solution (7.5%), sterile-filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
Guanosine 5′-triphosphate sodium salt solution, HPLC purified, aqueous solution for RNA polymerase transcription