Single-Cell RT-PCR, a Technique to Decipher the Electrical, Anatomical, and Genetic Determinants of Neuronal Diversity

Part of the Methods in Molecular Biology book series (MIMB, volume 1183)

Abstract

The patch-clamp technique has allowed for detailed studies on the electrical properties of neurons. Dye loading through patch pipettes enabled characterizing the morphological properties of the neurons. In addition, the patch-clamp technique also allows for harvesting mRNA from single cells to study gene expression at the single cell level (known as single-cell RT-PCR). The combination of these three approaches makes possible the study of the GEM profile of neurons (gene expression, electrophysiology, and morphology) using a single patch pipette and patch-clamp recording. This combination provides a powerful technique to investigate and correlate the neuron’s gene expression with its phenotype (electrical behavior and morphology). The harvesting and amplification of single cell mRNA for gene expression studies is a challenging task, especially for researchers with sparse or no training in molecular biology (seeNotes 1,2 and 5). Here we describe in detail the GEM profiling approach with special attention to the gene expression profiling.

Key words

Single-cell RT-PCR Multiplex PCR Ion channel Calcium binding protein Neuropeptide Morphology Electrophysiology 

Notes

Acknowledgements

The authors would like to thank the late Prof. Phil Goodman for helpful insights on the statistical approaches to the single-cell RT-PCR data analysis. We would like to thank Shaoling Ma, Claudia Herzberg, Raya Eilam, and Tal Hetzroni for their technical assistance and Dr Jesper Ryger for his useful comments. This work was supported by the National Alliance for Autism Research and a European Union grant.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Queens Medical Centre, School of Life SciencesUniversity of NottinghamNottinghamUK
  2. 2.Brain and Mind InstituteEPFLLausanneSwitzerland

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