Single-Cell RTPCR, a Technique to Decipher the Electrical, Anatomical, and Genetic Determinants of Neuronal Diversity
互联网
454
The patch-clamp technique has allowed detailed studies on the electrical properties of neurons. Dye loading through patch
pipettes has allowed characterizing the morphological properties of the neurons. In addition, the patch-clamp technique also
allows harvesting mRNA from single cells to study gene expression at the single-cell level (known as single-cell reverse transcription–polymerase
chain reaction [RT–PCR] [1-3]
). The combination of these three approaches allows determination of the Gene expression, Electrophysiology and Morphology
(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 study and correlate the neuron’s gene expression with its phenotype
(electrical behavior and morphology) (
4
–
7
)
. 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 (see
Notes 1
and 2
). Here, we describe in detail the GEM profiling approach with special attention to the gene expression profiling.