科研之友
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摘要
Objective: To investigate the CDR3 polymorphism of T cell receptor (TCR) in patients with vitiligo by high-throughput sequencing and immune repertoire analysis. Methods: A total of 45 patients with vitiligo were included in the case group, and 28 healthy volunteers were enrolled in the control group. Fasting venous blood collection was performed in both groups. Upon the total RNA extraction of cells, reverse transcription was carried out. Design and synthesis of downstream primers for TCR beta chain constant gene (TRBC) and specificity upstream primers for TCR beta chain variable gene (TRBV) gene family with common fluorescent labeling were conducted. Polymerase chain reaction was used for amplification of CDR3 in TRBV family. The CDR3 polymorphism and length of TRBV family were analyzed by means of a capillary electrophoresis genescan. Based on the analysis results, 8 samples in case group and 7 samples in control group were selected for cloning and sequencing of the TRBV6 family. The polymorphism, variable and joining gene expression as well as the composition, structure, and function of amino acid in CDR3 were also analyzed and the expression ratio of clonotype was detected. Results: Analysis of capillary electrophoresis genescan showed that compared with the control group, the CDR3 polymorphism in case group was significantly decreased, and the expressive abundance of TRBV20-1, TRBV7-9, and TRBV2 genes was the highest in the samples. The expression of TRBV7-9, TRBV25-1, TRBV4-1, TRBV12-5 showed significant differences, with the difference of TRBV7-9 being the most significant. The expressive abundance of clonotypes in different individuals differed greatly. There was no preference difference for amino acids within the groups, but there was an obvious difference in amino acid structure and function between the two groups. Analysis of a hyper-expressed clone found that the samples were highly heterogeneous, but the sequence similarity was very low, which proved that the expression difference of TRBV7-9 was the most obvious between the groups. Conclusion: The CDR3 polymorphism in patients with vitiligo was significantly increased.
- 出版日期2018
- 单位中山大学
