Gene Expression Analysis by Multiplex Single-Cell RT-PCR

Methods Mol Biol. 2019:1941:139-154. doi: 10.1007/978-1-4939-9077-1_10.

Abstract

Brain circuit assemblies comprise different cellular subpopulations that exhibit morphological, electrophysiological, and molecular diversity. Here we describe a protocol which, combined with whole-cell patch-clamp recording and morphological reconstruction, allows the transcriptomic analysis of the recorded cell. This protocol provides recipes on how to detect simultaneously the expression of 24 genes/markers at the single-cell level using polymerase chain reaction (PCR), how to design gene-specific probes, and how to validate them. This technique provides multiplexed expression data that cannot be easily obtained by other approaches such as immunological co-labeling.

Keywords: Cell type; Electrophysiology; Transcriptome; cDNA; mRNA.

MeSH terms

  • Cells, Cultured
  • Gene Expression Profiling / methods*
  • Humans
  • Neurons / cytology
  • Neurons / metabolism*
  • Patch-Clamp Techniques / methods*
  • RNA, Messenger / genetics*
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • Single-Cell Analysis / methods*

Substances

  • RNA, Messenger