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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 phosphate monobasic solution, BioUltra, 5 M in H2O
Pricing and availability is not currently available.
Sigma-Aldrich
Diethyl pyrocarbonate, 96% (NT)
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Sigma-Aldrich
Sodium phosphate monobasic, ReagentPlus®, ≥99.0%
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NBQX disodium salt hydrate, ≥98% (HPLC)
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Sucrose, ≥99.5% (GC)
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Guanosine 5′-triphosphate sodium salt solution, HPLC purified, aqueous solution for RNA polymerase transcription
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SR-95531, ≥98% (HPLC), powder
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Sucrose, BioUltra, for molecular biology, ≥99.5% (HPLC)
Pricing and availability is not currently available.
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Calcium chloride solution, BioUltra, for molecular biology, ~1 M in H2O
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Potassium D-gluconate, ≥99% (HPLC)
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DL-2-Amino-5-phosphonopentanoic acid, solid
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Sigma-Aldrich
Sodium bicarbonate, powder, BioReagent, for molecular biology, suitable for cell culture, suitable for insect cell culture
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D-(+)-Glucose, ≥99.5% (GC), BioXtra
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Adenosine 5′-triphosphate disodium salt solution, Crystalline ATP, HPLC purified, aqueous solution for RNA transcription
Pricing and availability is not currently available.
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Sodium bicarbonate solution, solution (7.5%), sterile-filtered, BioReagent, suitable for cell culture
Pricing and availability is not currently available.
Sigma-Aldrich
Hydrochloric acid solution, 1.0 N, BioReagent, suitable for cell culture
Pricing and availability is not currently available.
Supelco
JWH-018 5-Pentanoic acid beta-D-glucuronide solution, 100 μg/mL (Acetonitrile:Water (90:10) (v/v)), certified reference material, ampule of 1 mL
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Sigma-Aldrich
Acetic acid, glacial, ACS reagent, ≥99.7%
Pricing and availability is not currently available.
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Potassium chloride, ACS reagent, 99.0-100.5%
Pricing and availability is not currently available.
Sigma-Aldrich
Magnesium chloride solution, for molecular biology, 1.00 M±0.01 M
Pricing and availability is not currently available.