A Method for Simultaneously Detecting TCR Libraries of Multiple Trace Samples

Abstract

The invention discloses a method for simultaneously detecting multiple trace sample TCR (T cell receptor) repertoires. The method includes: taking a sample RNA (ribonucleic acid) as a template to synthesize a first-strand cDNA (complementary deoxyribonucleic acid), performing 5'-terminal RACE PCR, designing specific 5' RACE nested PCR primers and TCR alpha-beta gene specific primers, performing RT-PCR amplification to obtain alpha-beta gene amplification products of each sample TCR, mixing the multiple sample PCR products for sequencing, distinguishing different samples according to joining regions of the specific primers, and analyzing frequency distribution of V and J gene segments in each sample. The method for simultaneously detecting the multiple trace sample TCR repertoires has the advantages that on the premise that distinguishing of different samples is guaranteed, sequencing cost of the TCR repertoires is greatly reduced, and authenticity in frequency distribution of each family is guaranteed; establishment of disease entity RCR repertoires requiring a great quantity of samples is promoted.

Description

Technical field [0001] The invention belongs to the fields of biotechnology and clinical immunology, and specifically relates to a method for detecting T cell receptor (TCR) of multiple trace samples. Background technique [0002] T cells are the main components of lymphocytes. They have a variety of biological functions, such as direct killing of target cells, assisting or inhibiting B cells to produce antibodies, responding to specific antigens and mitogens, and producing cytokines, etc. A heroic fighter in the body to resist infection and tumor formation. T cells recognize pathogens and their toxins in the external environment and tumor antigens in the internal environment due to gene mutations through their surface receptor (T cell receptor, TCR) molecules. However, the antigens in the environment are ever-changing, so how can they recognize the ever-changing nature in nature antigen? The classical clonal selection theory believes that there are many lymphocyte clones in th...

AI Technical Summary

Problems solved by technology

②Due to the design of different primers for each family for RT-PCR amplification, the difference in the amplification efficiency of each TCR Vα and Vβ gene family itself will cause the deviation of the frequency distribution, which cannot objectively reflect the frequency distribution of each family

③ RT-PCR amplification of 58 families requires more cDNA templates and larger sample volume (usually at least 10ml of blood sample), and the operation process is cumbersome, time-consuming and labor-intensive, and prone to operational errors

Images