What are the steps involved in single-cell RNA sequencing?

There are 4 main steps involved in single-cell RNA sequencing: 

1. Cell isolation: Individual cells are isolated from a mixed population, using a variety of techniques such as fluorescence-activated cell sorting (FACS), magnetic-activated cell sorting (MACS), laser capture microdissection (LCM) or micromanipulation. 
2. Cell lysis and RNA capture: The isolated cells are lysed to release their RNA so that the maximum number of RNA molecules can be captured. Poly[T]-primers are often used to specifically capture polyadenylated mRNA molecules, which are then converted into complementary DNA (cDNA) by reverse transcription.
3. cDNA Amplification: The small amounts of cDNA are amplified using PCR or other methods. During this step, additional sequences, such as adaptors for next-generation sequencing (NGS) and unique molecular identifiers (UMIs), may be added to preserve information about the original mRNA molecules and their cellular origin.
4. Sequencing and Data Analysis: The amplified and tagged cDNA from each cell is pooled and sequenced using NGS. The resulting data is then analyzed using specialized bioinformatics tools to study gene expression at the single-cell level.