Single-cell transcriptome sequencing refers to the technology of reverse transcription, amplification, and sequencing of all RNA from a single cell, followed by bioinformatics, and it has developed rapidly. Due to its ability to interpret cell heterogeneity at the single cell level, single-cell transcriptome sequencing has been widely used in various fields of life science, especially in the research of diseases such as tumors. This article mainly introduces the process and development of single-cell transcriptome sequencing, and analyzes its application in oncology research such as tumor heterogeneity, tumor metastasis, tumor microenvironment, and anti-tumor drug development.