title: GSM6268650: SAMPLE1-CD4-TCR; Homo sapiens; OTHER
library_construction_protocol: T cells obtained from PBMCs stimulated by DCs  for 30 d were sorted by FACS. CD3+CD4+ T cells were collected, and 1×106/ml T cells were used in further 10x Genomics Chromium™ Single Cell V(D)J immune repertoire profiling performed and analysed by Novogene. Gel beads containing barcode information are mixed withsamples and enzymes and then combined with oil surfactants to form GEMs. The barcode sequence of the library built by V(D)J was connected to the switch oligo TSO sequence. The reverse transcription of mRNAs was reverse transcribed to the 5′ end by poly(T) primers, and CCC was added at the end to capture the cDNA. Transcription continued to add the barcode sequence to the 5′ end cDNA sequence. Nested PCR was performed using the universal primer of the 5′ adaptor and the continuous nested primers of the TCR constant region. This process was combined with the sequencing adaptor P5 to achieve the enrichment of the V(D)J region of the TCR. The enzymatic digestion method randomly interrupts the V(D)J cDNA. By controlling the interrupted sequences of different lengths, the entire V(D)J fragment cDNA sequence was covered, and the sequencing adapter P7 was added to perform PCR amplification. The main analysis content of single cell V(D)J included using Trimmomatic to trim the original data to obtain clean data and then using fastqc to evaluate the data quality of rawdata and cleandata. Then, TCR clone subtype analysis was performed by Novogene using the Cell Ranger software provided by 10x Genomics to perform each step of screening, contig stitching, and cloning type analysis and abundance analysis of samples, and VDJtools software was used to characterize V/J paired genes and CDR3 regions.