incorporates known sequences for the second round PCR primer sites, with sequencing adaptors and indices being subsequently incorporated. However, the high diversity at the primer sites of overlap sites in the first round PCR necessitates a large number of degenerated primers, thus predisposing to ...
Learn about our SMARTer approach to single T-cell sequencing that obtains alpha-beta pairing information.
Simple PCR amplification: a single primer pair for each TCR (alpha or beta) subunit per reaction UMI-based correction: removal of reads derived from PCR duplicates and sequencing errors Sensitive and specific clonotype detection: optimized cDNA library generation UDI...
sequencingT cell specificity is determined by the combinatorial association of specific variable (V), diversity (D), and junctional (J) regions. Clones of T cells (clonality) can occur, in the blood or in tissue, after proliferation of activated T cells. Determining clonality in mutation assays...
Antigen-specific T cells enrichment, bulk, and single-cell TCR sequencing PBMCs from patients were stimulated with HPV18E776-105peptide-pulsed autologous dendritic cells in vitro for 4 weeks. After stimulation, IFNγ+T cells were enriched by IFNγ Secretion Assay-Cell Enrichment and Detection Kit...
sequencing. Sequences of all primers are listed in Supplementary Table2. The first PCR reaction was performed using the Qiagen Multiplex PCR kit (Qiagen), in a total volume of 50 μL: 25 μL of Qiagen Multiplex PCR master mix, 10 μL of a mix of all primers (each primer at...
GeneStudio S5 next-generation sequencing systems. The assay kit provides a single pool of multiplex PCR primers, library reagents, and sample barcodes. Ion AmpliSeq libraries can be generated from research samples such as RNA extracted from whole blood, fresh-frozen tissue, or FACS-sorted cells....
第一轮PCR采用扩增过的双链cDNA为模板,正向引物与SMART sequence互补,同时包含Illumina Read 2序列(TCR Primer 1),反向引物与TCR-α和TCR-β的恒定区(即不变区)互补(TCR a/b Human Primer 1)。第二轮PCR以第一轮PCR产物为模板,采用的正向引物与第一轮PCR添加的Read 2 序列互补。反向引物与恒定区结合,位于...
upstream plasmid sequence and a reverse primer annealing to the TCRα chain C-region. In a second round PCR, the amplicon was extended to include adapters, which were used for another PCR extension with Oxford Nanopore Technology (ONT) barcode and sequencing adapters from the PCR Barcoding Kit ...
The DNA sequence of the TCRs from ELISA positive phage clones were obtained by sequencing using methods known to those skilled in the art. Example 6 Biacore Analysis of TCRs Obtained from the Library Method The affinity for antigen of the TCRs isolated from the library was determined by surfa...