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Each cell is unique, but most of the current research is directed at cell populations (such as whole tissue), ignoring the heterogeneity between cells. Single-cell sequencing is a new technology for simultaneous high-throughput sequencing of genomic, transcriptome, and epigenetic groups at the single cell level for up to tens of thousands of cells. Single-cell sequencing can more accurately analyze the difference between cells and cells, and realize the genetic detection from the "average value" to the "individual value", which is called the technology of gene sequencing 2.0 era. After the analysis, from the "individual value" to the "group value", it is possible to classify similar cell populations, count each type of cells and analyze their proportion and status in the population, which is a powerful clinical study and The platform for basic research.
At present, the single-cell RNA-Seq technology is the fastest growing single-cell sequencing. It can simultaneously analyze the gene expression status of thousands or even tens of thousands of individual cells, and has a wide range of research fields in cancer stem cells, tumor resistance mechanisms, and tumor immune microenvironment. application. At the end of 2018, single-cell technology with single-cell RNA-Seq as the main detection method was selected by Science as the top of the top ten scientific breakthroughs in 2018.
After more than ten years of development in the field of single cell sequencing, various technologies have emerged, including microfluidics, microdroplets, microplates, intracellular markers and other major methods. The single-cell RNA-Seq method mainly consists of a technology platform based on microdroplets or microfluidic chips. The former is represented by 10X Genomics' Chromium Single Cell System, which includes WaferGen's ICELL8 platform. 10X Genomics' single-cell RNA-Seq platform offers a wide range of applications in the industry due to its high throughput and high cell capture efficiency. 10X Genomics' single-cell RNA-Seq platform uses microfluidic sorting up to tens of thousands of single cells, encapsulating gel beads with primers, single-cell labeled barcodes, and reagents required for reaction in oil droplets. The oil droplets reverse-transcribe the RNA released by cell lysis to produce a barcode-bearing cDNA. After the oil droplets are destroyed, the cDNA is constructed into a library, and the library is analyzed by a second-generation sequencing platform. A large number of single-cell gene expression data can be obtained by one sequencing. . Another technique for single-cell studies is SMART-Seq, which can analyze the full length of transcriptome RNA and is used in many basic research fields. The disadvantage is that the flux is low and requires a lot of manual input.
Single-cell sequencing is a hot topic in current biomedical research. According to Pubmed's search for “single cell sequencing”, nearly 60% of articles have been published every year since 2011.
Single cell sequencing published articles
Pubmed search for "single cell sequencing"
Statistics on the number of articles published each year
Single-cell technology has helped pioneering basic and applied research in many fields; in 2018, Guo Guojun of Zhejiang University first described the single-cell map of mammals. Peking University's Deng Hongkui team used 10X Genomics technology to analyze chemically-induced stem cell reprogramming gene expression profiles. Single-cell technology has also been fruitful in the field of tumor heterogeneity research in 2018; the Navin team of MD Anderson in the United States has discovered the mechanism of drug resistance in triple-negative breast cancer, and the Harvard Medical School team revealed the heterogeneity of cancer cells in the head and neck. Cell transformation mechanism of cancer metastasis.
Single cell sequencing is the core technology platform for studying the tumor immune microenvironment. Peking University's Zhang Zemin team used single-cell sequencing technology to conduct pioneering research on the tumor microenvironment of liver cancer and colorectal cancer, and recently revealed the relationship between T cell subsets of lung cancer tumor microenvironment and patient prognosis. The Thienpont team in Belgium used 10X Genomics single-cell sequencing technology to reveal stromal cells from 52 lung cancer microenvironments. In addition to describing the tumor immune microenvironment, revealing microenvironmental subtypes and their relationship to tumor progression or prognosis, the application of single cell sequencing in immunotherapy clinical research has also begun to emerge. In November 2018, Cell published a work from the Broad Institute, using 10X Genomics technology to discover that melanoma promotes T cell rejection and tolerance to immunotherapy by inhibiting antigen presentation systems such as B2M and HLA-A/B/C. The mechanism; Cell reported a simultaneous study from Harvard Medical School, through single-cell sequencing, found that TCF7 + CD8 T cells can predict the efficacy of melanoma patients on immunotherapy. In addition, a team at the University of Washington discovered the mechanism of acquired drug resistance in 1018 Genomics single-cell sequencing platforms in 2018.
Single-cell sequencing will become an indispensable research tool to reveal tumor heterogeneity, targeted drug sensitivity and drug resistance mechanism, immunotherapy in different cancer populations and drug resistance mechanisms. Before the clinical transformation of single-cell sequencing technology, the standardization and automation of tissue separation into single cells, the integration of experimental and analytical procedures, and the interpretation of single-cell sequencing results continue to emerge new solutions. In the 2017 Single Cell Special Issue, Israeli scientist Ido Amit predicts that single-cell genetic analysis will become a routine clinical testing program in the next 10 years. We are waiting for it and are fully prepared for the arrival of the era of individualized single cell sequencing.