SMARTer Mouse TCR a/b Profiling Kit
Here we will discuss how to process TCR cDNA libraries obtained with SMARTer Mouse TCR a/b Profiling Kit.
Data libraries
This tutorial uses the data from the following publication: A T cell repertoire timestamp is at the core of responsiveness to CTLA-4 blockade Hagit Philip et al., , iScience, 2021 Feb; doi: 10.1016/j.isci.2021.102100
Samples were collected from 20 mice implanted with MC38 adenocarcinoma cells. The experiment contained 2 groups, of which the first group (5 animals with ids A - E) was dosed with 10 ml/kg/day of vehicle on day 1, 3 and 6 of the experiment. The second group (15 animals with ids F - T) was dosed with the investigational antibody anti-mCTLA4 (5/2.5 mg/kg/day at 1/3, 6). After treatment with the immune checkpoint inhibitor antibody anti-mCTLA4 blood samples for the preparation of PBMCs were taken at days 0,7,14 and 21. Tumors (where still visible) and spleens were collected immediately after euthanasia, and directly transferred to liquid nitrogen (snap-frozen samples). Total RNA was isolated from every sample and cDNA libraries were prepared using SMARTer Mouse TCR a/b Profiling Kit. TCR sequencing was performed on an Illumina Miseq sequencer using the 600-cycle Miseq reagent kit v3(Illumina) with pair-end, 2x300 base pair reads.
All data is available from SRA (PRJNA658348) using e.g. SRA Explorer.
Use aria2c for efficient download of the full dataset with the proper filenames:
download.sh
mkdir -p raw
aria2c -c -s 16 -x 16 -k 1M -j 8 -i download-list.txt
download-list.txt
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/021/SRR12487521/SRR12487521_1.fastq.gz
out=raw/S_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/022/SRR12487522/SRR12487522_1.fastq.gz
out=raw/S_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/023/SRR12487523/SRR12487523_1.fastq.gz
out=raw/S_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/024/SRR12487524/SRR12487524_1.fastq.gz
out=raw/S_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/025/SRR12487525/SRR12487525_1.fastq.gz
out=raw/S_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/026/SRR12487526/SRR12487526_1.fastq.gz
out=raw/I_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/027/SRR12487527/SRR12487527_1.fastq.gz
out=raw/I_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/028/SRR12487528/SRR12487528_1.fastq.gz
out=raw/I_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/029/SRR12487529/SRR12487529_1.fastq.gz
out=raw/I_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/030/SRR12487530/SRR12487530_1.fastq.gz
out=raw/I_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/031/SRR12487531/SRR12487531_1.fastq.gz
out=raw/T_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/032/SRR12487532/SRR12487532_1.fastq.gz
out=raw/J_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/033/SRR12487533/SRR12487533_1.fastq.gz
out=raw/J_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/034/SRR12487534/SRR12487534_1.fastq.gz
out=raw/J_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/035/SRR12487535/SRR12487535_1.fastq.gz
out=raw/J_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/036/SRR12487536/SRR12487536_1.fastq.gz
out=raw/J_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/037/SRR12487537/SRR12487537_1.fastq.gz
out=raw/A_t_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/038/SRR12487538/SRR12487538_1.fastq.gz
out=raw/A_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/039/SRR12487539/SRR12487539_1.fastq.gz
out=raw/A_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/040/SRR12487540/SRR12487540_1.fastq.gz
out=raw/A_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/041/SRR12487541/SRR12487541_1.fastq.gz
out=raw/M_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/042/SRR12487542/SRR12487542_1.fastq.gz
out=raw/T_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/043/SRR12487543/SRR12487543_1.fastq.gz
out=raw/M_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/044/SRR12487544/SRR12487544_1.fastq.gz
out=raw/M_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/045/SRR12487545/SRR12487545_1.fastq.gz
out=raw/M_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/046/SRR12487546/SRR12487546_1.fastq.gz
out=raw/M_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/047/SRR12487547/SRR12487547_1.fastq.gz
out=raw/G_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/048/SRR12487548/SRR12487548_1.fastq.gz
out=raw/G_t_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/049/SRR12487549/SRR12487549_1.fastq.gz
out=raw/G_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/050/SRR12487550/SRR12487550_1.fastq.gz
out=raw/G_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/051/SRR12487551/SRR12487551_1.fastq.gz
out=raw/G_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/052/SRR12487552/SRR12487552_1.fastq.gz
out=raw/G_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/053/SRR12487553/SRR12487553_1.fastq.gz
out=raw/T_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/054/SRR12487554/SRR12487554_1.fastq.gz
out=raw/O_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/055/SRR12487555/SRR12487555_1.fastq.gz
out=raw/O_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/056/SRR12487556/SRR12487556_1.fastq.gz
out=raw/O_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/057/SRR12487557/SRR12487557_1.fastq.gz
out=raw/O_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/058/SRR12487558/SRR12487558_1.fastq.gz
out=raw/O_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/059/SRR12487559/SRR12487559_1.fastq.gz
out=raw/R_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/060/SRR12487560/SRR12487560_1.fastq.gz
out=raw/R_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/061/SRR12487561/SRR12487561_1.fastq.gz
out=raw/R_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/062/SRR12487562/SRR12487562_1.fastq.gz
out=raw/R_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/063/SRR12487563/SRR12487563_1.fastq.gz
out=raw/R_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/064/SRR12487564/SRR12487564_1.fastq.gz
out=raw/T_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/065/SRR12487565/SRR12487565_1.fastq.gz
out=raw/H_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/066/SRR12487566/SRR12487566_1.fastq.gz
out=raw/H_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/067/SRR12487567/SRR12487567_1.fastq.gz
out=raw/H_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/068/SRR12487568/SRR12487568_1.fastq.gz
out=raw/H_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/069/SRR12487569/SRR12487569_1.fastq.gz
out=raw/H_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/070/SRR12487570/SRR12487570_1.fastq.gz
out=raw/P_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/071/SRR12487571/SRR12487571_1.fastq.gz
out=raw/P_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/072/SRR12487572/SRR12487572_1.fastq.gz
out=raw/P_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/073/SRR12487573/SRR12487573_1.fastq.gz
out=raw/P_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/074/SRR12487574/SRR12487574_1.fastq.gz
out=raw/P_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/075/SRR12487575/SRR12487575_1.fastq.gz
out=raw/T_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/076/SRR12487576/SRR12487576_1.fastq.gz
out=raw/E_t_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/077/SRR12487577/SRR12487577_1.fastq.gz
out=raw/E_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/078/SRR12487578/SRR12487578_1.fastq.gz
out=raw/E_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/079/SRR12487579/SRR12487579_1.fastq.gz
out=raw/E_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/080/SRR12487580/SRR12487580_1.fastq.gz
out=raw/E_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/081/SRR12487581/SRR12487581_1.fastq.gz
out=raw/E_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/082/SRR12487582/SRR12487582_1.fastq.gz
out=raw/D_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/083/SRR12487583/SRR12487583_1.fastq.gz
out=raw/D_t_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/084/SRR12487584/SRR12487584_1.fastq.gz
out=raw/D_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/085/SRR12487585/SRR12487585_1.fastq.gz
out=raw/D_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/086/SRR12487586/SRR12487586_1.fastq.gz
out=raw/Q_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/087/SRR12487587/SRR12487587_1.fastq.gz
out=raw/D_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/088/SRR12487588/SRR12487588_1.fastq.gz
out=raw/D_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/089/SRR12487589/SRR12487589_1.fastq.gz
out=raw/K_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/090/SRR12487590/SRR12487590_1.fastq.gz
out=raw/K_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/091/SRR12487591/SRR12487591_1.fastq.gz
out=raw/K_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/092/SRR12487592/SRR12487592_1.fastq.gz
out=raw/K_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/093/SRR12487593/SRR12487593_1.fastq.gz
out=raw/L_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/094/SRR12487594/SRR12487594_1.fastq.gz
out=raw/L_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/095/SRR12487595/SRR12487595_1.fastq.gz
out=raw/L_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/096/SRR12487596/SRR12487596_1.fastq.gz
out=raw/L_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/097/SRR12487597/SRR12487597_1.fastq.gz
out=raw/Q_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/098/SRR12487598/SRR12487598_1.fastq.gz
out=raw/L_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/099/SRR12487599/SRR12487599_1.fastq.gz
out=raw/F_t_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/000/SRR12487600/SRR12487600_1.fastq.gz
out=raw/F_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/001/SRR12487601/SRR12487601_1.fastq.gz
out=raw/F_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/002/SRR12487602/SRR12487602_1.fastq.gz
out=raw/F_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/003/SRR12487603/SRR12487603_1.fastq.gz
out=raw/F_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/004/SRR12487604/SRR12487604_1.fastq.gz
out=raw/F_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/005/SRR12487605/SRR12487605_1.fastq.gz
out=raw/N_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/006/SRR12487606/SRR12487606_1.fastq.gz
out=raw/N_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/007/SRR12487607/SRR12487607_1.fastq.gz
out=raw/N_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/008/SRR12487608/SRR12487608_1.fastq.gz
out=raw/Q_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/009/SRR12487609/SRR12487609_1.fastq.gz
out=raw/N_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/010/SRR12487610/SRR12487610_1.fastq.gz
out=raw/N_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/011/SRR12487611/SRR12487611_1.fastq.gz
out=raw/C_s_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/012/SRR12487612/SRR12487612_1.fastq.gz
out=raw/C_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/013/SRR12487613/SRR12487613_1.fastq.gz
out=raw/C_21_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/014/SRR12487614/SRR12487614_1.fastq.gz
out=raw/C_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/015/SRR12487615/SRR12487615_1.fastq.gz
out=raw/C_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/016/SRR12487616/SRR12487616_1.fastq.gz
out=raw/B_7_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/017/SRR12487617/SRR12487617_1.fastq.gz
out=raw/B_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/018/SRR12487618/SRR12487618_1.fastq.gz
out=raw/B_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/019/SRR12487619/SRR12487619_1.fastq.gz
out=raw/Q_14_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/020/SRR12487620/SRR12487620_1.fastq.gz
out=raw/Q_0_R1.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/021/SRR12487521/SRR12487521_2.fastq.gz
out=raw/S_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/022/SRR12487522/SRR12487522_2.fastq.gz
out=raw/S_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/023/SRR12487523/SRR12487523_2.fastq.gz
out=raw/S_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/024/SRR12487524/SRR12487524_2.fastq.gz
out=raw/S_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/025/SRR12487525/SRR12487525_2.fastq.gz
out=raw/S_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/026/SRR12487526/SRR12487526_2.fastq.gz
out=raw/I_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/027/SRR12487527/SRR12487527_2.fastq.gz
out=raw/I_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/028/SRR12487528/SRR12487528_2.fastq.gz
out=raw/I_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/029/SRR12487529/SRR12487529_2.fastq.gz
out=raw/I_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/030/SRR12487530/SRR12487530_2.fastq.gz
out=raw/I_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/031/SRR12487531/SRR12487531_2.fastq.gz
out=raw/T_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/032/SRR12487532/SRR12487532_2.fastq.gz
out=raw/J_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/033/SRR12487533/SRR12487533_2.fastq.gz
out=raw/J_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/034/SRR12487534/SRR12487534_2.fastq.gz
out=raw/J_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/035/SRR12487535/SRR12487535_2.fastq.gz
out=raw/J_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/036/SRR12487536/SRR12487536_2.fastq.gz
out=raw/J_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/037/SRR12487537/SRR12487537_2.fastq.gz
out=raw/A_t_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/038/SRR12487538/SRR12487538_2.fastq.gz
out=raw/A_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/039/SRR12487539/SRR12487539_2.fastq.gz
out=raw/A_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/040/SRR12487540/SRR12487540_2.fastq.gz
out=raw/A_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/041/SRR12487541/SRR12487541_2.fastq.gz
out=raw/M_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/042/SRR12487542/SRR12487542_2.fastq.gz
out=raw/T_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/043/SRR12487543/SRR12487543_2.fastq.gz
out=raw/M_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/044/SRR12487544/SRR12487544_2.fastq.gz
out=raw/M_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/045/SRR12487545/SRR12487545_2.fastq.gz
out=raw/M_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/046/SRR12487546/SRR12487546_2.fastq.gz
out=raw/M_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/047/SRR12487547/SRR12487547_2.fastq.gz
out=raw/G_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/048/SRR12487548/SRR12487548_2.fastq.gz
out=raw/G_t_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/049/SRR12487549/SRR12487549_2.fastq.gz
out=raw/G_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/050/SRR12487550/SRR12487550_2.fastq.gz
out=raw/G_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/051/SRR12487551/SRR12487551_2.fastq.gz
out=raw/G_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/052/SRR12487552/SRR12487552_2.fastq.gz
out=raw/G_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/053/SRR12487553/SRR12487553_2.fastq.gz
out=raw/T_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/054/SRR12487554/SRR12487554_2.fastq.gz
out=raw/O_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/055/SRR12487555/SRR12487555_2.fastq.gz
out=raw/O_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/056/SRR12487556/SRR12487556_2.fastq.gz
out=raw/O_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/057/SRR12487557/SRR12487557_2.fastq.gz
out=raw/O_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/058/SRR12487558/SRR12487558_2.fastq.gz
out=raw/O_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/059/SRR12487559/SRR12487559_2.fastq.gz
out=raw/R_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/060/SRR12487560/SRR12487560_2.fastq.gz
out=raw/R_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/061/SRR12487561/SRR12487561_2.fastq.gz
out=raw/R_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/062/SRR12487562/SRR12487562_2.fastq.gz
out=raw/R_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/063/SRR12487563/SRR12487563_2.fastq.gz
out=raw/R_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/064/SRR12487564/SRR12487564_2.fastq.gz
out=raw/T_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/065/SRR12487565/SRR12487565_2.fastq.gz
out=raw/H_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/066/SRR12487566/SRR12487566_2.fastq.gz
out=raw/H_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/067/SRR12487567/SRR12487567_2.fastq.gz
out=raw/H_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/068/SRR12487568/SRR12487568_2.fastq.gz
out=raw/H_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/069/SRR12487569/SRR12487569_2.fastq.gz
out=raw/H_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/070/SRR12487570/SRR12487570_2.fastq.gz
out=raw/P_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/071/SRR12487571/SRR12487571_2.fastq.gz
out=raw/P_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/072/SRR12487572/SRR12487572_2.fastq.gz
out=raw/P_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/073/SRR12487573/SRR12487573_2.fastq.gz
out=raw/P_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/074/SRR12487574/SRR12487574_2.fastq.gz
out=raw/P_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/075/SRR12487575/SRR12487575_2.fastq.gz
out=raw/T_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/076/SRR12487576/SRR12487576_2.fastq.gz
out=raw/E_t_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/077/SRR12487577/SRR12487577_2.fastq.gz
out=raw/E_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/078/SRR12487578/SRR12487578_2.fastq.gz
out=raw/E_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/079/SRR12487579/SRR12487579_2.fastq.gz
out=raw/E_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/080/SRR12487580/SRR12487580_2.fastq.gz
out=raw/E_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/081/SRR12487581/SRR12487581_2.fastq.gz
out=raw/E_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/082/SRR12487582/SRR12487582_2.fastq.gz
out=raw/D_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/083/SRR12487583/SRR12487583_2.fastq.gz
out=raw/D_t_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/084/SRR12487584/SRR12487584_2.fastq.gz
out=raw/D_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/085/SRR12487585/SRR12487585_2.fastq.gz
out=raw/D_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/086/SRR12487586/SRR12487586_2.fastq.gz
out=raw/Q_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/087/SRR12487587/SRR12487587_2.fastq.gz
out=raw/D_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/088/SRR12487588/SRR12487588_2.fastq.gz
out=raw/D_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/089/SRR12487589/SRR12487589_2.fastq.gz
out=raw/K_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/090/SRR12487590/SRR12487590_2.fastq.gz
out=raw/K_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/091/SRR12487591/SRR12487591_2.fastq.gz
out=raw/K_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/092/SRR12487592/SRR12487592_2.fastq.gz
out=raw/K_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/093/SRR12487593/SRR12487593_2.fastq.gz
out=raw/L_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/094/SRR12487594/SRR12487594_2.fastq.gz
out=raw/L_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/095/SRR12487595/SRR12487595_2.fastq.gz
out=raw/L_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/096/SRR12487596/SRR12487596_2.fastq.gz
out=raw/L_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/097/SRR12487597/SRR12487597_2.fastq.gz
out=raw/Q_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/098/SRR12487598/SRR12487598_2.fastq.gz
out=raw/L_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/099/SRR12487599/SRR12487599_2.fastq.gz
out=raw/F_t_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/000/SRR12487600/SRR12487600_2.fastq.gz
out=raw/F_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/001/SRR12487601/SRR12487601_2.fastq.gz
out=raw/F_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/002/SRR12487602/SRR12487602_2.fastq.gz
out=raw/F_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/003/SRR12487603/SRR12487603_2.fastq.gz
out=raw/F_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/004/SRR12487604/SRR12487604_2.fastq.gz
out=raw/F_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/005/SRR12487605/SRR12487605_2.fastq.gz
out=raw/N_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/006/SRR12487606/SRR12487606_2.fastq.gz
out=raw/N_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/007/SRR12487607/SRR12487607_2.fastq.gz
out=raw/N_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/008/SRR12487608/SRR12487608_2.fastq.gz
out=raw/Q_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/009/SRR12487609/SRR12487609_2.fastq.gz
out=raw/N_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/010/SRR12487610/SRR12487610_2.fastq.gz
out=raw/N_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/011/SRR12487611/SRR12487611_2.fastq.gz
out=raw/C_s_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/012/SRR12487612/SRR12487612_2.fastq.gz
out=raw/C_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/013/SRR12487613/SRR12487613_2.fastq.gz
out=raw/C_21_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/014/SRR12487614/SRR12487614_2.fastq.gz
out=raw/C_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/015/SRR12487615/SRR12487615_2.fastq.gz
out=raw/C_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/016/SRR12487616/SRR12487616_2.fastq.gz
out=raw/B_7_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/017/SRR12487617/SRR12487617_2.fastq.gz
out=raw/B_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/018/SRR12487618/SRR12487618_2.fastq.gz
out=raw/B_0_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/019/SRR12487619/SRR12487619_2.fastq.gz
out=raw/Q_14_R2.fastq.gz
ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR124/020/SRR12487620/SRR12487620_2.fastq.gz
out=raw/Q_0_R2.fastq.gz
The project contains 100 FASTQ file pairs. For the purpose of this tutorial we assume that all fastq files are stored in fastq/ folder. Each file name encodes the information about mouse id and PMBC sample time-point (0,7,14,21 days) or tissue (s - spleen, t -tumor) - S - mouce id, experiment group - s - spleen tissue
The structure of the library is shown on the picture bellow.
Upstream analysis
MiXCR has a dedicated preset for this protocol, thus analysing the data is as easy as:
mixcr analyze takara-mouse-rna-tcr-smarter \
raw/S_s_R1.fastq.gz \
raw/S_s_R2.fastq.gz \
results/S_s
One might also use GNU Parallel to process all samples at once:
#!/usr/bin/env bash
mkdir -p results
ls /raw/*R1* |
parallel -j 2 --line-buffer \
"mixcr analyze takara-mouse-rna-tcr-smarter \
{} \
{=s:R1:R2:=} \
{=s:.*/:results/:;s:_R.*::=}"
Under the hood the command above actually executes the following pipeline:
Under the hood pipeline:
Under the hood the command above actually executes the following pipeline:
align
Alignment of raw sequencing reads against reference database of V-, D-, J- and C- gene segments.
mixcr align \
--species mmu \
-p generic-amplicon \
-OvParameters.geneFeatureToAlign="VTranscriptWithout5UTRWithP" \
-OvParameters.parameters.floatingLeftBound=false \
-OjParameters.parameters.floatingRightBound=false \
--report results/S_s.report \
--json-report results/S_s.json \
raw/S_s_R1.fastq.gz \
raw/S_s_R2.fastq.gz \
results/S_s.vdjca
Option --report is specified here explicitly.
--species mmu- determines the organism species (mmu for Mus Musculus).
-p generic-amplicon- a default preset of MiXCR parameters for amplicon data.
-OvParameters.geneFeatureToAlign="VTranscriptWithout5UTRWithP"- Sets a V gene feature to align. Check gene features for more info.
-OvParameters.parameters.floatingLeftBound=false- Results in a global alignment algorithm for V gene left bound. We use it because we don't have any primers covering V gene coding sequence.
-OjParameters.parameters.floatingRightBound=false- Results in a global alignment algorithm for J gene right bound, because reverse primers are located in C-gene region.
assemble
Assembles alignments into clonotypes and applies several layers of errors correction(ex. quality-dependent correction for sequencing errors, PCR-error correction by clustering). Check mixcr assemble for more information.
mixcr assemble \
-OassemblingFeatures="CDR3"
-OseparateByJ=true \
-OseparateByV=true \
--report results/S_s.report \
--json-report results/S_s.json \
results/S_s.vdjca \
results/S_s.clns
Options --report and --json-report are specified here explicitly so that the report files will be appended with assembly report.
-OassemblingFeatures=CDR3- By default
takara-mmu-tcr-cdr3preset assembles clones byCDR3sequence.
export
Exports clonotypes from .clns file into human-readable tables.
mixcr exportClones \
-c TRA \
results/S_s.clns \
results/S_s.clonotypes.TRA.txt
-с <chain>- defines a specific chain to be exported.
After executing the command above for every sample we will have a following set of output files:
# human-readable reports
S_s.report
# raw alignments (highly compressed binary file)
S_s.vdjca
# TCRα & TCRβ CDR3 clonotypes (highly compressed binary file)
S_s.clns
# TCRα & TCRβ CDR3 clonotypes exported in tab-delimited txt
S_s.TRB.txt
S_s.TRB.txt
While .clns file holds all data and is used for downstream analysis, the output .txt clonotype table will contain exhaustive information about each clonotype as well:
See first 100 records from S_s clonotype table
| cloneId | cloneCount | cloneFraction | targetSequences | targetQualities | allVHitsWithScore | allDHitsWithScore | allJHitsWithScore | allCHitsWithScore | allVAlignments | allDAlignments | allJAlignments | allCAlignments | nSeqFR1 | minQualFR1 | nSeqCDR1 | minQualCDR1 | nSeqFR2 | minQualFR2 | nSeqCDR2 | minQualCDR2 | nSeqFR3 | minQualFR3 | nSeqCDR3 | minQualCDR3 | nSeqFR4 | minQualFR4 | aaSeqFR1 | aaSeqCDR1 | aaSeqFR2 | aaSeqCDR2 | aaSeqFR3 | aaSeqCDR3 | aaSeqFR4 | refPoints |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 0 | 194 | 0.0056568 | TGTGTGGTGGGCGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1349),TRAV11D00(1349) | nan | TRAJ18*00(314.5) | nan | 267 | 280 | 300 | 0 | 13 | 65.0;267 | 280 | 300 | 0 | 13 | 65.0 | nan | 22 | 55 | 86 | 12 | 45 | 165.0 | nan | nan | nan | nan | nan | |||
| 1 | 135 | 0.00393643 | TGTGTGGTGGGGGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1363),TRAV11D00(1363) | nan | TRAJ18*00(316.6) | nan | 267 | 278 | 300 | 0 | 11 | 55.0;267 | 278 | 300 | 0 | 11 | 55.0 | nan | 22 | 55 | 86 | 12 | 45 | 165.0 | nan | nan | nan | nan | nan | |||
| 2 | 104 | 0.00303251 | TGTGTGGTGGGAGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1380.9),TRAV11D00(1380.9) | nan | TRAJ18*00(320.8) | nan | 267 | 278 | 300 | 0 | 11 | 55.0;267 | 278 | 300 | 0 | 11 | 55.0 | nan | 21 | 55 | 86 | 11 | 45 | 170.0 | nan | nan | nan | nan | nan | |||
| 3 | 66 | 0.00192448 | TGTGCAGTGAATAGGGGTAGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7N-6*00(1062.4) | nan | TRAJ31*00(270.8) | nan | 264 | 274 | 297 | 0 | 10 | 50.0 | nan | 24 | 46 | 77 | 17 | 39 | 110.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 4 | 53 | 0.00154541 | TGTGTGGTGGGTGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1382.6),TRAV11D00(1382.6) | nan | TRAJ18*00(316.2) | nan | 267 | 278 | 300 | 0 | 11 | 55.0;267 | 278 | 300 | 0 | 11 | 55.0 | nan | 22 | 55 | 86 | 12 | 45 | 165.0 | nan | nan | nan | nan | nan | |||
| 5 | 34 | 0.000991398 | TGTGCAGCAAGTAGGGATTATAACCAGGGGAAGCTTATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14N-3*00(1280.9) | nan | TRAJ23*00(280.2) | nan | 267 | 279 | 300 | 0 | 12 | 60.0 | nan | 23 | 49 | 80 | 16 | 42 | 130.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 6 | 33 | 0.000962239 | TGTGCTATGAGAGAGGGCAGCAACAACAGACTTACTTTG | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV16D-DV11*00(1370.6) | nan | TRAJ7*00(260.7) | nan | 273 | 291 | 310 | 0 | 18 | 90.0 | nan | 26 | 48 | 79 | 17 | 39 | 110.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 7 | 32 | 0.000933081 | TGTGCTTTGGAACATAATGCAGGTGCCAAGCTCACATTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(1047.1) | nan | TRAJ39*00(283.3) | nan | 258 | 271 | 291 | 0 | 13 | 65.0 | nan | 26 | 52 | 83 | 13 | 39 | 130.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 8 | 29 | 0.000845604 | TGTGCCAGCAGCCCCGGGACAGGGGTTGGAAATACGCTCTATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV5*00(1343.2) | TRBD1*00(60) | TRBJ1-3*00(228) | nan | 267 | 280 | 304 | 0 | 13 | 65.0 | 10 | 22 | 36 | 13 | 25 | 60.0 | 23 | 42 | 70 | 26 | 45 | 95.0 | nan | nan | nan | nan | nan | |||
| 9 | 27 | 0.000787287 | TGTGCAGCAAGTGAGGGAGGCAGAGCTCTGATATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14N-3*00(1179.1) | nan | TRAJ15*00(274) | nan | 267 | 280 | 300 | 0 | 13 | 65.0 | nan | 26 | 49 | 80 | 13 | 36 | 115.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 10 | 25 | 0.000728969 | TGTGCTTCCTTGAATAGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(1018.5) | nan | TRAJ31*00(276.7) | nan | 258 | 265 | 291 | 0 | 7 | 35.0 | nan | 21 | 46 | 77 | 11 | 36 | 125.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 11 | 25 | 0.000728969 | TGTGCAGTGAGGTATAATACAGGAAACTACAAATACGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-5*00(1327) | nan | TRAJ40*00(304.1) | nan | 264 | 275 | 297 | 0 | 11 | 55.0 | nan | 22 | 50 | 81 | 14 | 42 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 12 | 25 | 0.000728969 | TGTGCAGCAAGCATATGGGATTATAACCAGGGGAAGCTTATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1000(1410.5),TRAV10D00(1380.8) | nan | TRAJ23*00(283.3) | nan | 267 | 281 | 300 | 0 | 14 | 70.0;267 | 281 | 300 | 0 | 14 | 70.0 | nan | 23 | 49 | 80 | 19 | 45 | 130.0 | nan | nan | nan | nan | nan | |||
| 13 | 23 | 0.000670652 | TGTGCCAGCTCTCTCGTGGGGGCAAACTCCGACTACACCTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV12-1*00(1217.9) | TRBD2*00(25) | TRBJ1-2*00(234.5) | nan | 270 | 286 | 305 | 0 | 16 | 80.0 | 19 | 24 | 42 | 16 | 21 | 25.0 | 19 | 40 | 68 | 21 | 42 | 105.0 | nan | nan | nan | nan | nan | |||
| 14 | 21 | 0.000612334 | TGTGCTATGAGAGAGGGCTACAGCAACAACAGACTTACTTTG | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV16D-DV1100(1376.7),TRAV16N00(1362.7) | nan | TRAJ7*00(272.7) | nan | 273 | 291 | 310 | 0 | 18 | 90.0;273 | 291 | 310 | 0 | 18 | 90.0 | nan | 20 | 48 | 79 | 14 | 42 | SA22G | 126.0 | nan | nan | nan | nan | nan | ||
| 15 | 20 | 0.000583175 | TGTGTGGTGGGCGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGFGGGGGGGGGGGGGGGGGGG | TRAV11D*00(1532.3) | nan | TRAJ18*00(307.2) | nan | 267 | 280 | 300 | 0 | 13 | 65.0 | nan | 22 | 55 | 86 | 12 | 45 | 165.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 16 | 20 | 0.000583175 | TGTGCTATGGAAAGTGATGGAAATGAGAAAATAACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-4-DV7*00(772.4) | nan | TRAJ48*00(270) | nan | 258 | 273 | 291 | 0 | 15 | SC270A | 61.0 | nan | 27 | 50 | 81 | 16 | 39 | 115.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 17 | 20 | 0.000583175 | TGTGCAGCAAGTTCTAACAGTGCAGGGAACAAGCTAACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14N-3*00(1298.5) | nan | TRAJ17*00(299.5) | nan | 267 | 279 | 300 | 0 | 12 | 60.0 | nan | 23 | 52 | 83 | 13 | 42 | 145.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 18 | 20 | 0.000583175 | TGTGGTGCTAGACTGGGGGGGCGCGGGTATGCTGAGCAGTTCTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV20*00(1367.4) | TRBD2*00(65) | TRBJ2-1*00(225.8) | nan | 279 | 290 | 313 | 0 | 11 | 55.0 | 17 | 30 | 42 | 11 | 24 | 65.0 | 24 | 42 | 70 | 27 | 45 | 90.0 | nan | nan | nan | nan | nan | |||
| 19 | 19 | 0.000554017 | TGTGCAGTGAGCGACAATACAGAAGGTGCAGATAGACTCACCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-6*00(895.1) | nan | TRAJ45*00(326.1) | nan | 264 | 276 | 297 | 0 | 12 | 60.0 | nan | 20 | 49 | 80 | 16 | 45 | 145.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 20 | 18 | 0.000524858 | TGTGCTTCCATGTCTAATTACAACGTGCTTTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(734.3) | nan | TRAJ21*00(295) | nan | 258 | 264 | 291 | 0 | 6 | 30.0 | nan | 18 | 46 | 77 | 8 | 36 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 21 | 18 | 0.000524858 | TGTGCTGCCCAAGGAGGGTCTGCGAAGCTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV4-4-DV10*00(1181.1) | nan | TRAJ57*00(282.3) | nan | 258 | 266 | 291 | 0 | 8 | 40.0 | nan | 25 | 52 | 83 | 9 | 36 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 22 | 17 | 0.000495699 | TGTGCTTTGGAGCGTGGAGGCAGCAATTACAAACTGACATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(851.6) | nan | TRAJ53*00(303.5) | nan | 258 | 273 | 291 | 0 | 15 | SA269G | 61.0 | nan | 26 | 55 | 86 | 13 | 42 | 145.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 23 | 17 | 0.000495699 | TGTGCTACTCCTATAACAGGCAATACCGGAAAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV8-1*00(1333.3) | nan | TRAJ37*00(303.5) | nan | 264 | 273 | 297 | 0 | 9 | 45.0 | nan | 18 | 49 | 80 | 11 | 42 | 155.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 24 | 17 | 0.000495699 | TGTGCAGTGAGCATGAGGGTTATGGGAGCAGTGGCAACAAGCTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-5*00(1195.4) | nan | TRAJ32*00(309.1) | nan | 264 | 279 | 297 | 0 | 15 | 75.0 | nan | 24 | 55 | 86 | 19 | 50 | 155.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 25 | 17 | 0.000495699 | TGCTCCAGCAGCCCCGGGAGGGACACAGAAGTCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV23*00(628.2) | TRBD100(35),TRBD200(35) | TRBJ1-1*00(218.3) | nan | 273 | 284 | 310 | 0 | 11 | 55.0 | 9 | 16 | 36 | 12 | 19 | 35.0;11 | 18 | 42 | 12 | 19 | 35.0 | 23 | 40 | 68 | 22 | 39 | ||||
| 26 | 16 | 0.00046654 | TGTGCTTTTGGAGGCAGCAATTACAAACTGACATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(318.3) | nan | TRAJ53*00(289.1) | nan | 258 | 269 | 291 | 0 | 12 | I263T | 43.0 | nan | 27 | 55 | 86 | 8 | 36 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 27 | 16 | 0.00046654 | TGTGCTTTGGGTTATAACACCAATACAGGCAAATTAACCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(305.8) | nan | TRAJ27*00(329.7) | nan | 258 | 268 | 291 | 0 | 10 | 50.0 | nan | 11 | 48 | 79 | 6 | 42 | DT15 | 168.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 28 | 16 | 0.00046654 | TGTGCTATGGAACAAGAAAGAACACCAATACAGGCAAATTAACCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(391) | nan | TRAJ27*00(307.8) | nan | 258 | 271 | 291 | 0 | 13 | 65.0 | nan | 21 | 48 | 79 | 20 | 47 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 29 | 16 | 0.00046654 | TGTGCTACTGATGGAGGGACTGGAGGCTATAAAGTGGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV8-1*00(1394.9) | nan | TRAJ12*00(287.4) | nan | 264 | 277 | 297 | 0 | 13 | 65.0 | nan | 21 | 48 | 79 | 15 | 42 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 30 | 16 | 0.00046654 | TGTGCAGCAAGGACTAATTACAACGTGCTTTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1000(1384.8),TRAV10D00(1356.8) | nan | TRAJ21*00(264.8) | nan | 267 | 278 | 300 | 0 | 11 | 55.0;267 | 278 | 300 | 0 | 11 | 55.0 | nan | 23 | 46 | 77 | 13 | 36 | 115.0 | nan | nan | nan | nan | nan | |||
| 31 | 16 | 0.00046654 | TGCACCTGCAGTGCAGAGGGACAGGGGTGGGATGCAGAAACGCTGTATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV1*00(1107.6) | TRBD1*00(50) | TRBJ2-3*00(222.5) | nan | 270 | 287 | 307 | 0 | 17 | 85.0 | 12 | 22 | 36 | 17 | 27 | 50.0 | 22 | 41 | 69 | 32 | 51 | 95.0 | nan | nan | nan | nan | nan | |||
| 32 | 15 | 0.000437382 | TGTGCTATGGATTCCGGGACTGGAGGCTATAAAGTGGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(330.2),TRAV13N-100(330.2) | nan | TRAJ12*00(287.5) | nan | 258 | 269 | 291 | 0 | 11 | 55.0;258 | 269 | 291 | 0 | 11 | 55.0 | nan | 21 | 48 | 79 | 15 | 42 | 135.0 | nan | nan | nan | nan | nan | |||
| 33 | 15 | 0.000437382 | TGTGCTATGGATGACACCAATACAGGCAAATTAACCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(389.7) | nan | TRAJ27*00(285.1) | nan | 258 | 269 | 291 | 0 | 11 | 55.0 | nan | 19 | 48 | 79 | 10 | 39 | SA21G | 131.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 34 | 15 | 0.000437382 | TGTGCTATGGAACCTAATTACAACGTGCTTTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(291.9),TRAV13N-100(291.9) | nan | TRAJ21*00(275.9) | nan | 258 | 271 | 291 | 0 | 13 | 65.0;258 | 271 | 291 | 0 | 13 | 65.0 | nan | 23 | 46 | 77 | 13 | 36 | 115.0 | nan | nan | nan | nan | nan | |||
| 35 | 15 | 0.000437382 | TGTGCAGCAAGCTTTAGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14-1*00(1353.3) | nan | TRAJ31*00(265) | nan | 267 | 278 | 300 | 0 | 11 | 55.0 | nan | 24 | 46 | 77 | 14 | 36 | 110.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 36 | 15 | 0.000437382 | TGTGCAGCAAGCATGAACAGTGCAGGGAACAAGCTAACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-200(980.4),TRAV7D-200(980.4) | nan | TRAJ17*00(289) | nan | 264 | 279 | 297 | 0 | 15 | 75.0;264 | 279 | 297 | 0 | 15 | 75.0 | nan | 25 | 52 | 83 | 15 | 42 | 135.0 | nan | nan | nan | nan | nan | |||
| 37 | 14 | 0.000408223 | TGTGCTGCTGAGGCCAACTATGGAAATGAGAAAATAACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV4-4-DV10*00(1078.3) | nan | TRAJ48*00(292.2) | nan | 258 | 273 | 291 | 0 | 15 | 75.0 | nan | 20 | 50 | 81 | 12 | 42 | 150.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 38 | 14 | 0.000408223 | TGTGCTATGGCGGGCACAAATGCTTACAAAGTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(1087.2) | nan | TRAJ30*00(289.8) | nan | 258 | 268 | 291 | 0 | 10 | 50.0 | nan | 23 | 48 | 79 | 14 | 39 | 125.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 39 | 14 | 0.000408223 | TGTGCTATAGACTACAGAAGGTGCAGATAGACTCACCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-2*00(1339.2) | nan | TRAJ45*00(284.2) | nan | 258 | 269 | 291 | 0 | 11 | 55.0 | nan | 21 | 49 | 80 | 12 | 40 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 40 | 14 | 0.000408223 | TGTGCAGTGTACACAAATGCTTACAAAGTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-6*00(772.7) | nan | TRAJ30*00(314.8) | nan | 264 | 273 | 297 | 0 | 9 | 45.0 | nan | 13 | 48 | 79 | 0 | 36 | DT17I20GI22T | 134.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 41 | 14 | 0.000408223 | TGTGCAGTGAGATCTGGCAGCTGGCAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-5*00(1385.9) | nan | TRAJ22*00(269.8) | nan | 264 | 275 | 297 | 0 | 11 | 55.0 | nan | 25 | 49 | 80 | 12 | 36 | 120.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 42 | 14 | 0.000408223 | TGCACCTGCAGTGCAGCCGGGACTTTTAACCAAGACACCCAGTACTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV1*00(1088.8) | TRBD200(40),TRBD100(35) | TRBJ2-5*00(245.4) | nan | 270 | 286 | 307 | 0 | 16 | 80.0 | 12 | 20 | 42 | 16 | 24 | 40.0;10 | 17 | 36 | 16 | 23 | 35.0 | 18 | 41 | 69 | 25 | 48 | ||||
| 43 | 13 | 0.000379064 | TGTGTGGTTAGGAATACCAACAAAGTCGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1236),TRAV11D00(1236) | nan | TRAJ34*00(260) | nan | 267 | 275 | 300 | 0 | 8 | 40.0;267 | 275 | 300 | 0 | 8 | 40.0 | nan | 26 | 47 | 78 | 12 | 33 | 105.0 | nan | nan | nan | nan | nan | |||
| 44 | 13 | 0.000379064 | TGTGCTATGGGGGCTATAACAGGCAATACCGGAAAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-1*00(419.1) | nan | TRAJ37*00(310) | nan | 258 | 268 | 291 | 0 | 10 | 50.0 | nan | 18 | 49 | 80 | 14 | 45 | 155.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 45 | 13 | 0.000379064 | TGTGCTATGGGGATGAATTATAACCAGGGGAAGCTTATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(503.5),TRAV13N-100(502.3) | nan | TRAJ23*00(328.2) | nan | 258 | 268 | 291 | 0 | 10 | 50.0;258 | 268 | 291 | 0 | 10 | 50.0 | nan | 19 | 49 | 80 | 12 | 42 | 150.0 | nan | nan | nan | nan | nan | |||
| 46 | 13 | 0.000379064 | TGTGCTATAGACCTGGACACAAATGCTTACAAAGTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-200(1029.8),TRAV13N-200(1029.8) | nan | TRAJ30*00(285.7) | nan | 258 | 269 | 291 | 0 | 11 | 55.0;258 | 269 | 291 | 0 | 11 | 55.0 | nan | 21 | 48 | 79 | 15 | 42 | 135.0 | nan | nan | nan | nan | nan | |||
| 47 | 13 | 0.000379064 | TGTGCCAGCTCTCTCGACTGGGGGGACACCGGGCAGCTCTACTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV12-2*00(1099.8) | TRBD2*00(40) | TRBJ2-2*00(247.5) | nan | 270 | 287 | 305 | 0 | 17 | 85.0 | 18 | 26 | 42 | 17 | 25 | 40.0 | 23 | 43 | 71 | 25 | 45 | 100.0 | nan | nan | nan | nan | nan | |||
| 48 | 13 | 0.000379064 | TGTGCCAGCGGTGAATACTGGGGGACTAGTCAAAACACCTTGTACTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV13-2*00(1240.5) | TRBD2*00(35) | TRBJ2-4*00(251.6) | nan | 270 | 284 | 306 | 0 | 14 | 70.0 | 17 | 24 | 42 | 16 | 23 | 35.0 | 16 | 41 | 69 | 23 | 48 | 125.0 | nan | nan | nan | nan | nan | |||
| 49 | 13 | 0.000379064 | TGTGCAGCAAAGAGTGGCAGTGGTGGAAAACTCACTTTG | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1000(1270.1),TRAV10D00(1246.8) | nan | TRAJ44*00(276.3) | nan | 267 | 277 | 300 | 0 | 10 | 50.0;267 | 277 | 300 | 0 | 10 | 50.0 | nan | 25 | 50 | 81 | 14 | 39 | 125.0 | nan | nan | nan | nan | nan | |||
| 50 | 12 | 0.000349905 | TGTGTGGTGGGAGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV11D*00(1547.7) | nan | TRAJ18*00(317.7) | nan | 267 | 278 | 300 | 0 | 11 | 55.0 | nan | 21 | 55 | 86 | 11 | 45 | 170.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 51 | 12 | 0.000349905 | TGTGCTTTGGGCCGTTATGGGAGCAGTGGCAACAAGCTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(1041) | nan | TRAJ32*00(305.5) | nan | 258 | 274 | 291 | 0 | 16 | SA268GSA269C | 52.0 | nan | 24 | 55 | 86 | 14 | 45 | 155.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 52 | 12 | 0.000349905 | TGTGCTATCCTTGACACAAATGCTTACAAAGTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(391.4) | nan | TRAJ30*00(290.3) | nan | 258 | 266 | 291 | 0 | 8 | 40.0 | nan | 20 | 48 | 79 | 11 | 39 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 53 | 12 | 0.000349905 | TGTGCTATGGTCCTAAATGCTTACAAAGTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(318.9) | nan | TRAJ30*00(262.2) | nan | 258 | 268 | 291 | 0 | 10 | 50.0 | nan | 26 | 48 | 79 | 14 | 36 | 110.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 54 | 12 | 0.000349905 | TGTGCTATGGAACGGGGAGGGTCTGCGAAGCTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(368.5),TRAV13N-100(368.5) | nan | TRAJ57*00(272.5) | nan | 258 | 272 | 291 | 0 | 14 | 70.0;258 | 272 | 291 | 0 | 14 | 70.0 | nan | 28 | 52 | 83 | 15 | 39 | 120.0 | nan | nan | nan | nan | nan | |||
| 55 | 12 | 0.000349905 | TGTGCAGTGTCCACTACTGGAGGCAATAATAAGCTGACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-5*00(1348.5) | nan | TRAJ56*00(298.7) | nan | 264 | 273 | 297 | 0 | 9 | 45.0 | nan | 23 | 52 | 83 | 13 | 42 | 145.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 56 | 12 | 0.000349905 | TGTGCAGCCCAGACAGGCTTTGCAAGTGCGCTGACATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7D-400(1165.7),TRAV7-400(1151.7) | nan | TRAJ35*00(317.9) | nan | 264 | 272 | 301 | 0 | 8 | 40.0;264 | 272 | 301 | 0 | 8 | 40.0 | nan | 18 | 51 | 85 | 6 | 39 | SG20C | 151.0 | nan | nan | nan | nan | nan | ||
| 57 | 12 | 0.000349905 | TGTGCAGCAAGTGACTACAGCAACAACAGACTTACTTTG | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14N-3*00(1182.3) | nan | TRAJ7*00(288.4) | nan | 267 | 284 | 300 | 0 | 16 | DC280 | 68.0 | nan | 21 | 48 | 79 | 12 | 39 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 58 | 11 | 0.000320746 | TGTGCTTTGAGTGACCGCCAGGGAGGCAGAGCTCTGATATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV12-2*00(1046.2) | nan | TRAJ15*00(275.3) | nan | 267 | 281 | 301 | 0 | 14 | 70.0 | nan | 24 | 49 | 80 | 17 | 42 | 125.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 59 | 11 | 0.000320746 | TGTGCTTTAACAGGCAATACCGGAAAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(1035.5) | nan | TRAJ37*00(296.2) | nan | 258 | 266 | 291 | 0 | 8 | 40.0 | nan | 20 | 49 | 80 | 7 | 36 | 145.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 60 | 11 | 0.000320746 | TGTGCTGCCTACCAGGGAGGCAGAGCTCTGATATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(266.8),TRAV13N-100(266.8) | nan | TRAJ15*00(303.4) | nan | 258 | 264 | 291 | 0 | 6 | 30.0;258 | 264 | 291 | 0 | 6 | 30.0 | nan | 19 | 49 | 80 | 6 | 36 | 150.0 | nan | nan | nan | nan | nan | |||
| 61 | 11 | 0.000320746 | TGTGCTATGGTAGCAAATTCTGGGACTTACCAGAGGTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(395.1),TRAV13N-100(395.1) | nan | TRAJ13*00(287.2) | nan | 258 | 268 | 291 | 0 | 10 | 50.0;258 | 268 | 291 | 0 | 10 | 50.0 | nan | 19 | 46 | 77 | 12 | 39 | 135.0 | nan | nan | nan | nan | nan | |||
| 62 | 11 | 0.000320746 | TGTGCTATGGGGCCCTCTTCTGGCAGCTGGCAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(375.8),TRAV13N-100(375.8) | nan | TRAJ22*00(282.4) | nan | 258 | 268 | 291 | 0 | 10 | 50.0;258 | 268 | 291 | 0 | 10 | 50.0 | nan | 22 | 49 | 80 | 15 | 42 | 135.0 | nan | nan | nan | nan | nan | |||
| 63 | 11 | 0.000320746 | TGTGCTATGGAACACGGCAGGGAGGCAGAGCTCTGATATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(336.8),TRAV13N-100(336.8) | nan | TRAJ15*00(294.1) | nan | 258 | 271 | 291 | 0 | 13 | 65.0;258 | 271 | 291 | 0 | 13 | 65.0 | nan | 25 | 49 | 80 | 17 | 41 | 120.0 | nan | nan | nan | nan | nan | |||
| 64 | 11 | 0.000320746 | TGTGCCAGCAGCCAAGACGGGGGGGCCCACTCCTATGAACAGTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV5*00(1313.2) | TRBD200(40),TRBD100(36) | TRBJ2-7*00(235) | nan | 267 | 284 | 304 | 0 | 17 | 85.0 | 20 | 28 | 42 | 18 | 26 | 40.0;18 | 28 | 36 | 18 | 28 | SC23G | 36.0 | 20 | 39 | 67 | 29 | 48 | |||
| 65 | 11 | 0.000320746 | TGTGCAGTGAGCATTCGTGGAGCTAACACTGGAAAGCTCACGTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7N-6*00(788.5) | nan | TRAJ52*00(289.1) | nan | 264 | 278 | 297 | 0 | 14 | 70.0 | nan | 27 | 55 | 86 | 17 | 45 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 66 | 11 | 0.000320746 | TGTGCAGCAAGTGCAGAGACTGGAGGCTATAAAGTGGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14D-200(1282.1),TRAV14N-200(1282.1) | nan | TRAJ12*00(280) | nan | 267 | 282 | 300 | 0 | 15 | 75.0;267 | 282 | 300 | 0 | 15 | 75.0 | nan | 23 | 48 | 79 | 17 | 42 | 125.0 | nan | nan | nan | nan | nan | |||
| 67 | 11 | 0.000320746 | TGTGCAGATAGCAACTATCAGTTGATCTGG | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-1*00(1269.2) | nan | TRAJ33*00(245.7) | nan | 264 | 277 | 298 | 0 | 13 | ST271ASG272T | 37.0 | nan | 22 | 46 | 77 | 6 | 30 | 120.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 68 | 11 | 0.000320746 | CCCCACATAATGCAGGTGCCAAGCTCACATTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7D-500(1196.3),TRAV7N-500(1196.3) | nan | TRAJ39*00(282.5) | nan | 264 | 264 | 297 | 0 | 0 | 0.0;264 | 264 | 297 | 0 | 0 | 0.0 | nan | 26 | 52 | 83 | 6 | 32 | 130.0 | nan | nan | nan | nan | nan | |||
| 69 | 10 | 0.000291588 | TGTCCGTCTAATTACAACGTGCTTTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(1006.3) | nan | TRAJ21*00(280) | nan | 258 | 261 | 291 | 0 | 3 | 15.0 | nan | 21 | 46 | 77 | 5 | 30 | 125.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 70 | 10 | 0.000291588 | TGTGCTTTGGAACGGGGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(371) | nan | TRAJ31*00(255) | nan | 258 | 272 | 291 | 0 | 14 | 70.0 | nan | 26 | 46 | 77 | 16 | 36 | 100.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 71 | 10 | 0.000291588 | TGTGCTATGGTCATGAATTATAACCAGGGGAAGCTTATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(1060.6),TRAV13N-100(1050.1) | nan | TRAJ23*00(311.5) | nan | 258 | 268 | 291 | 0 | 10 | 50.0;258 | 268 | 291 | 0 | 10 | 50.0 | nan | 17 | 49 | 80 | 10 | 42 | 160.0 | nan | nan | nan | nan | nan | |||
| 72 | 10 | 0.000291588 | TGTGCTATGGAATACCTACCAGGCACTGGGAGTAACAGGCTCACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(521.3),TRAV13N-100(521.3) | nan | TRAJ28*00(316.9) | nan | 258 | 270 | 291 | 0 | 12 | 60.0;258 | 270 | 291 | 0 | 12 | 60.0 | nan | 21 | 54 | 85 | 15 | 48 | 165.0 | nan | nan | nan | nan | nan | |||
| 73 | 10 | 0.000291588 | TGTGCTATGGAACCTGGTCTTCTGGCAGCTGGCAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(309.6) | nan | TRAJ22*00(288.6) | nan | 258 | 271 | 291 | 0 | 13 | 65.0 | nan | 22 | 49 | 80 | 17 | 44 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 74 | 10 | 0.000291588 | TGTGCCAGCAGTTTGACGGCCAACACCGGGCAGCTCTACTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV14*00(1433.4) | nan | TRBJ2-2*00(234.8) | nan | 273 | 287 | 309 | 0 | 14 | 70.0 | nan | 19 | 43 | 71 | 18 | 42 | SA21C | 106.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 75 | 10 | 0.000291588 | TGTGCCAGCAGTGATAAGGGGAACTATGCTGAGCAGTTCTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV13-1*00(1341.9) | TRBD1*00(25) | TRBJ2-1*00(239.4) | nan | 270 | 285 | 306 | 0 | 15 | 75.0 | 17 | 22 | 36 | 16 | 21 | 25.0 | 21 | 42 | 70 | 21 | 42 | 105.0 | nan | nan | nan | nan | nan | |||
| 76 | 10 | 0.000291588 | TGTGCGGCCAATACAGGCAAATTAACCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(280.1) | nan | TRAJ27*00(270) | nan | 258 | 263 | 291 | 0 | 5 | 25.0 | nan | 25 | 48 | 79 | 7 | 30 | 115.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 77 | 10 | 0.000291588 | TGTGCAGTGAGCCTTGTTAATACAGGAAACTACAAATACGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-6*00(723.2) | nan | TRAJ40*00(322.4) | nan | 264 | 276 | 297 | 0 | 12 | 60.0 | nan | 20 | 50 | 81 | 15 | 45 | 150.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 78 | 10 | 0.000291588 | TGTGCAGTGAGCATGGACTCGGGATACAACAAACTCACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7D-500(1367.6),TRAV7N-500(1367.6) | nan | TRAJ11*00(290) | nan | 264 | 279 | 297 | 0 | 15 | 75.0;264 | 279 | 297 | 0 | 15 | 75.0 | nan | 14 | 48 | 79 | 8 | 42 | DT17I22G | 141.0 | nan | nan | nan | nan | nan | ||
| 79 | 10 | 0.000291588 | TGTGCAGCCGCAGGCTTTGCAAGTGCGCTGACATTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV7-200(1335.4),TRAV7D-200(1335.4) | nan | TRAJ35*00(276.6) | nan | 264 | 282 | 297 | 0 | 17 | DA272SA273CST277G | 45.0;264 | 282 | 297 | 0 | 17 | DA272SA273CST277G | 45.0 | nan | 25 | 51 | 85 | 10 | 36 | 130.0 | nan | nan | nan | nan | nan | |
| 80 | 10 | 0.000291588 | TGTGCAGCAAGTGTGTCTAATTACAACGTGCTTTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14D-100(1387.6),TRAV14N-100(1387.6) | nan | TRAJ21*00(281.9) | nan | 267 | 280 | 300 | 0 | 13 | 65.0;267 | 280 | 300 | 0 | 13 | 65.0 | nan | 20 | 46 | 77 | 13 | 39 | 130.0 | nan | nan | nan | nan | nan | |||
| 81 | 10 | 0.000291588 | TGTGCAGCAAGTGACCCTTCTGGCAGCTGGCAACTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14N-3*00(330.2) | nan | TRAJ22*00(263.8) | nan | 267 | 289 | 300 | 0 | 22 | I280ASA281CDG285 | 67.0 | nan | 23 | 49 | 80 | 16 | 42 | 130.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | |
| 82 | 10 | 0.000291588 | TGTGCAGCAAGTAGGGGGGGCAGTGCAGGGAACAAGCTAACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV14D-100(1369.9),TRAV14N-100(1369.9) | nan | TRAJ17*00(278.6) | nan | 267 | 279 | 300 | 0 | 12 | 60.0;267 | 279 | 300 | 0 | 12 | 60.0 | nan | 27 | 52 | 83 | 20 | 45 | 125.0 | nan | nan | nan | nan | nan | |||
| 83 | 10 | 0.000291588 | TGCGCAGTCACCCTAACTATGCCCAGGGATTAACCTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV3D-300(661.9),TRAV3N-300(661.9) | nan | TRAJ26*00(277.6) | nan | 267 | 277 | 300 | 0 | 10 | 50.0;267 | 277 | 300 | 0 | 10 | 50.0 | nan | 24 | 49 | 80 | 13 | 38 | 125.0 | nan | nan | nan | nan | nan | |||
| 84 | 9 | 0.000262429 | TGTGTGGTGGCCGATAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1301.4),TRAV11D00(1301.4) | nan | TRAJ18*00(316.9) | nan | 267 | 280 | 300 | 0 | 13 | SG277C | 51.0;267 | 280 | 300 | 0 | 13 | SG277C | 51.0 | nan | 22 | 55 | 86 | 12 | 45 | 165.0 | nan | nan | nan | nan | nan | |
| 85 | 9 | 0.000262429 | TGTGTGGTGGGCGACAGAGGTTCAGCCTTAGGGAGGCTGCATTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV1100(1410),TRAV11D00(1410) | nan | TRAJ18*00(306) | nan | 267 | 280 | 300 | 0 | 13 | 65.0;267 | 280 | 300 | 0 | 13 | 65.0 | nan | 22 | 55 | 86 | 12 | 45 | ST24C | 151.0 | nan | nan | nan | nan | nan | ||
| 86 | 9 | 0.000262429 | TGTGCTTTGGGGGGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(384.7) | nan | TRAJ31*00(247.2) | nan | 258 | 268 | 291 | 0 | 10 | 50.0 | nan | 26 | 46 | 77 | 13 | 33 | 100.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 87 | 9 | 0.000262429 | TGTGCTTTGGAACGTGACTCGGGATACAACAAACTCACTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(760.4) | nan | TRAJ11*00(317.3) | nan | 258 | 273 | 291 | 0 | 15 | 75.0 | nan | 20 | 48 | 79 | 14 | 42 | 140.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 88 | 9 | 0.000262429 | TGTGCTCTCTGGGAGCTGGATAGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV15-1-DV6-1*00(1175.3) | nan | TRAJ31*00(266.9) | nan | 273 | 292 | 312 | 0 | 19 | 95.0 | nan | 23 | 46 | 77 | 19 | 42 | 115.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 89 | 9 | 0.000262429 | TGTGCTCTGGGTGATGATAGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV6-600(1355.6),TRAV6D-600(1355.6) | nan | TRAJ31*00(259) | nan | 273 | 288 | 307 | 0 | 15 | 75.0;273 | 288 | 307 | 0 | 15 | 75.0 | nan | 23 | 46 | 77 | 16 | 39 | 115.0 | nan | nan | nan | nan | nan | |||
| 90 | 9 | 0.000262429 | TGTGCTCTGAGCTCCGTTAATACAGGAAACTACAAATACGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV6-7-DV9*00(1252.2) | nan | TRAJ40*00(341.8) | nan | 267 | 278 | 301 | 0 | 11 | 55.0 | nan | 19 | 50 | 81 | 14 | 45 | 155.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 91 | 9 | 0.000262429 | TGTGCTCCCGACACAAATGCTTACAAAGTCATCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13-1*00(1211.1) | nan | TRAJ30*00(288) | nan | 258 | 264 | 291 | 0 | 6 | 30.0 | nan | 21 | 48 | 79 | 9 | 36 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 92 | 9 | 0.000262429 | TGTGCTGCTTACAACGTGCTTTACTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(237) | nan | TRAJ21*00(244.8) | nan | 258 | 264 | 291 | 0 | 6 | 30.0 | nan | 27 | 46 | 77 | 8 | 27 | 95.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 93 | 9 | 0.000262429 | TGTGCTGCTGAGGCGGGAGGCTATAAAGTGGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV4D-300(1355.3),TRAV4N-300(1355.3) | nan | TRAJ12*00(258.3) | nan | 258 | 272 | 291 | 0 | 14 | 70.0;258 | 272 | 291 | 0 | 14 | 70.0 | nan | 27 | 48 | 79 | 15 | 36 | 105.0 | nan | nan | nan | nan | nan | |||
| 94 | 9 | 0.000262429 | TGTGCTATGGCGGCTTCAAGTGGCCAGAAGCTGGTTTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(411.9),TRAV13N-100(411.9) | nan | TRAJ16*00(268.5) | nan | 258 | 268 | 291 | 0 | 10 | 50.0;258 | 268 | 291 | 0 | 10 | 50.0 | nan | 24 | 50 | 81 | 13 | 39 | 130.0 | nan | nan | nan | nan | nan | |||
| 95 | 9 | 0.000262429 | TGTGCTATGGACAAAAATACTGGAGGACTAAGTGGTAAATTAACATTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-300(1073.5),TRAV13D-300(1057.8) | nan | TRAJ2*00(307.4) | nan | 264 | 275 | 297 | 0 | 11 | 55.0;264 | 275 | 297 | 0 | 11 | 55.0 | nan | 22 | 55 | 86 | 15 | 48 | 165.0 | nan | nan | nan | nan | nan | |||
| 96 | 9 | 0.000262429 | TGTGCTATGGAACGTAATACAGGAAACTACAAATACGTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13D-100(299.7),TRAV13N-100(299.7) | nan | TRAJ40*00(295) | nan | 258 | 273 | 291 | 0 | 15 | 75.0;258 | 273 | 291 | 0 | 15 | 75.0 | nan | 17 | 50 | 81 | 10 | 42 | DT22 | 148.0 | nan | nan | nan | nan | nan | ||
| 97 | 9 | 0.000262429 | TGTGCTATGGAACGAATAATGCAGGTGCCAAGCTCACATTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(1043.7) | nan | TRAJ39*00(288.4) | nan | 258 | 272 | 291 | 0 | 14 | 70.0 | nan | 25 | 52 | 83 | 14 | 41 | 135.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 98 | 9 | 0.000262429 | TGTGCTATAGGGTAGCAATAACAGAATCTTCTTT | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRAV13N-4*00(264.4) | nan | TRAJ31*00(265) | nan | 258 | 266 | 291 | 0 | 8 | 40.0 | nan | 24 | 46 | 77 | 12 | 34 | 110.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | ||
| 99 | 9 | 0.000262429 | TGTGCCTGGAGTACTACAGGGGGAATCTATGCTGAGCAGTTCTTC | GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG | TRBV31*00(1236.1) | nan | TRBJ2-1*00(211.7) | nan | 270 | 292 | 303 | 0 | 20 | DC282DG283SC287A | 62.0 | nan | 23 | 42 | 70 | 26 | 45 | 95.0 | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan | nan |
Quality control
Now when we have processed all samples, we can proceed to quality control. First thing that we want to look at is the alignment quality. This can be easily done with the following command:
mixcr exportQc align results/*.clns figs/alignQc.pdf
From the plot above we see, that only about 50% of reads from every sample have been successfully aligned. The major reason why the alignment failed is that a lot of reads lack TCR sequences. In that case, if we want to dig a bit deeper into the issue we can realign one of the samples and save not-aligned reads into separate files for manual inspection. Let's pick one of the samples, ex. S_7, and realign it.
mkdir -p debug
mixcr align -f \
--species mmu \
-OvParameters.geneFeatureToAlign="VTranscriptWithout5UTRWithP" \
-OvParameters.parameters.floatingLeftBound=false \
-OjParameters.parameters.floatingRightBound=false \
--not-aligned-R1 debug/S_s_R1.fastq \
--not-aligned-R2 debug/S_s_R2.fastq \
--report debug/S_s.report \
--json-report debug/S_s.json \
raw/S_s_R1.fastq.gz \
raw/S_s_R2.fastq.gz \
debug/S_s.vdjca
Now one can use BLAST to determine the origin of the reads in S_7_notAligned_R1.fastq and S_7_notAligned_R2.fastq. In this particular case BLAST showed that major part of the non-aligned reads aligns with DNA sequence in between TCR gene segments which may be consequences of DNA contamination.
Finally, we can look at the chain distribution among all clones.
mixcr exportQc chainUsage results/*.clns figs/chainUsage.pdf
On the plot bellow we see, despite some TRB dominance, both (TCR alpha and TCR beta) chains equally present in each sample.
Full-length clonotype assembly
SMARTer Mouse TCR a/b Profiling Kit allows to recover a broader TCR receptor sequence then just CDR3 region. Since this protocol is based on 5'RACE method, the full V-gene sequence should present. Reverse sequence specific primers are located in C-gene region, thus the read fully covers J-gene.
The longest possible assembling feature for this protocol is "{FR1Begin:FR4End}".
MiXCR has a specific preset to obtain full-length TCR clones with SMARTer Mouse TCR a/b Profiling protocol:
mixcr analyze takara-mouse-rna-tcr-smarter \
raw/S_s_R1.fastq.gz \
raw/S_s_R2.fastq.gz \
results/S_s.vdjca
The mixcr assemble step in this preset differs from the one above in the following manner:
mixcr assemble \
-OassemblingFeatures="{FR1Begin:FR4End}" \
--report results/S_s.report \
--json-report results/S_s.json \
results/S_s.vdjca \
results/S_s.clns
-OassemblingFeatures="{FR1Begin:FR4End}"- sets the assembling feature to the region which starts from
FR1Beginand ends at the end ofFR4.
Notice that we do not use -OseparateByV=true and -OseparateByJ=true in this case because assembling feature already covers full V and J sequences, thus in case several clones have identical CDR3 they will still be separated.
Reports
Finally, MiXCR provides a very convenient way to look at the reports generated at ech step. Every .vdjca, .clns and .clna file holds all the reports for every MiXCR function that has been applied to this sample. E.g. in our case .clns file contains reports for mixcr align and mixcr assemble. To output this report use mixcr exportReports as shown bellow. Note --json parameter will output a JSON-formatted report.
mixcr exportReports \
results/S_s.clns \
results/S_s.report.txt
mixcr exportReports \
--json \
results/S_s.clns \
results/S_s.report.json
Show report file
============== Align Report ==============
Input file(s): /raw/PRJNA658348_TakaraMouseTCR/S_s_R1.fastq.gz,/raw/PRJNA658348_TakaraMouseTCR/S_s_R2.fastq.gz
Output file(s): results/S_s.vdjca
Version: ; built=Mon Sep 26 10:55:18 CEST 2022; rev=8c998df1ab; lib=repseqio.v2.0
Command line arguments: --report results/S_s.align.report.txt --json-report results/S_s.align.report.json --preset local:takara-smarter-mouse-tcr-cdr3 +limitInput 100000 /raw/PRJNA658348_TakaraMouseTCR/S_s_R1.fastq.gz /raw/PRJNA658348_TakaraMouseTCR/S_s_R2.fastq.gz results/S_s.vdjca
Analysis time: 0ns
Total sequencing reads: 100000
Successfully aligned reads: 48077 (48.08%)
Paired-end alignment conflicts eliminated: 6461 (6.46%)
Alignment failed, no hits (not TCR/IG?): 35335 (35.34%)
Alignment failed because of absence of V hits: 10019 (10.02%)
Alignment failed because of absence of J hits: 6103 (6.1%)
No target with both V and J alignments: 400 (0.4%)
Alignment failed because of low total score: 66 (0.07%)
Overlapped: 72072 (72.07%)
Overlapped and aligned: 34890 (34.89%)
Alignment-aided overlaps: 1306 (3.74%)
Overlapped and not aligned: 37182 (37.18%)
No CDR3 parts alignments, percent of successfully aligned: 135 (0.28%)
Partial aligned reads, percent of successfully aligned: 1862 (3.87%)
V gene chimeras: 118 (0.12%)
J gene chimeras: 25 (0.03%)
TRA chains: 30220 (62.86%)
TRA non-functional: 4116 (13.62%)
TRB chains: 17849 (37.13%)
TRB non-functional: 1382 (7.74%)
TRD chains: 8 (0.02%)
TRD non-functional: 2 (25%)
Realigned with forced non-floating bound: 58468 (58.47%)
Realigned with forced non-floating right bound in left read: 7379 (7.38%)
Realigned with forced non-floating left bound in right read: 7379 (7.38%)
============== Assemble Report ==============
Input file(s): results/S_s.vdjca
Output file(s): results/S_s.clns
Version: ; built=Mon Sep 26 10:55:18 CEST 2022; rev=8c998df1ab; lib=repseqio.v2.0
Command line arguments: --report results/S_s.assemble.report.txt --json-report results/S_s.assemble.report.json results/S_s.vdjca results/S_s.clns
Analysis time: 0ns
Final clonotype count: 25533
Average number of reads per clonotype: 1.34
Reads used in clonotypes, percent of total: 34295 (34.3%)
Reads used in clonotypes before clustering, percent of total: 34336 (34.34%)
Number of reads used as a core, percent of used: 33568 (97.76%)
Mapped low quality reads, percent of used: 768 (2.24%)
Reads clustered in PCR error correction, percent of used: 41 (0.12%)
Reads pre-clustered due to the similar VJC-lists, percent of used: 85 (0.25%)
Reads dropped due to the lack of a clone sequence, percent of total: 2655 (2.65%)
Reads dropped due to a too short clonal sequence, percent of total: 0 (0%)
Reads dropped due to low quality, percent of total: 45 (0.04%)
Reads dropped due to failed mapping, percent of total: 10984 (10.98%)
Reads dropped with low quality clones, percent of total: 77 (0.08%)
Clonotypes eliminated by PCR error correction: 22
Clonotypes dropped as low quality: 45
Clonotypes pre-clustered due to the similar VJC-lists: 72
TRA chains: 15308 (59.95%)
TRA non-functional: 2060 (13.46%)
TRB chains: 10224 (40.04%)
TRB non-functional: 635 (6.21%)
TRD chains: 1 (0%)
TRD non-functional: 1 (100%)
{
"type": "alignerReport",
"commandLine": "--report results/S_s.align.report.txt --json-report results/S_s.align.report.json --preset local:takara-smarter-mouse-tcr-cdr3 +limitInput 100000 /raw/PRJNA658348_TakaraMouseTCR/S_s_R1.fastq.gz /raw/PRJNA658348_TakaraMouseTCR/S_s_R2.fastq.gz results/S_s.vdjca",
"inputFiles": [
"/raw/PRJNA658348_TakaraMouseTCR/S_s_R1.fastq.gz",
"/raw/PRJNA658348_TakaraMouseTCR/S_s_R2.fastq.gz"
],
"outputFiles": [
"results/S_s.vdjca"
],
"version": "; built=Mon Sep 26 10:55:18 CEST 2022; rev=8c998df1ab; lib=repseqio.v2.0",
"trimmingReport": null,
"totalReadsProcessed": 100000,
"aligned": 48077,
"notAligned": 51923,
"notAlignedReasons": {
"NoVHits": 10019,
"LowTotalScore": 66,
"NoCDR3Parts": 0,
"VAndJOnDifferentTargets": 400,
"NoBarcode": 0,
"NoJHits": 6103,
"NoHits": 35335
},
"chimeras": 0,
"overlapped": 72072,
"alignmentAidedOverlaps": 1306,
"overlappedAligned": 34890,
"overlappedNotAligned": 37182,
"pairedEndAlignmentConflicts": 6461,
"vChimeras": 118,
"jChimeras": 25,
"chainUsage": {
"type": "chainUsage",
"chimeras": 0,
"total": 48077,
"chains": {
"TRA": {
"total": 30220,
"nonFunctional": 4116,
"isOOF": 3698,
"hasStops": 418
},
"TRB": {
"total": 17849,
"nonFunctional": 1382,
"isOOF": 1035,
"hasStops": 347
},
"TRD": {
"total": 8,
"nonFunctional": 2,
"isOOF": 2,
"hasStops": 0
}
}
},
"realignedWithForcedNonFloatingBound": 58468,
"realignedWithForcedNonFloatingRightBoundInLeftRead": 7379,
"realignedWithForcedNonFloatingLeftBoundInRightRead": 7379,
"noCDR3PartsAlignments": 135,
"partialAlignments": 1862,
"tagReport": null
}
{
"type": "assemblerReport",
"commandLine": "--report results/S_s.assemble.report.txt --json-report results/S_s.assemble.report.json results/S_s.vdjca results/S_s.clns",
"inputFiles": [
"results/S_s.vdjca"
],
"outputFiles": [
"results/S_s.clns"
],
"version": "; built=Mon Sep 26 10:55:18 CEST 2022; rev=8c998df1ab; lib=repseqio.v2.0",
"preCloneAssemblerReport": null,
"totalReadsProcessed": 100000,
"initialClonesCreated": 25672,
"readsDroppedNoTargetSequence": 2655,
"readsDroppedTooShortClonalSequence": 0,
"readsDroppedLowQuality": 25,
"coreReads": 33568,
"readsDroppedFailedMapping": 10984,
"lowQualityRescued": 768,
"clonesClustered": 22,
"readsClustered": 41,
"clones": 25533,
"clonesDroppedAsLowQuality": 45,
"clonesPreClustered": 72,
"readsPreClustered": 85,
"readsInClones": 34295,
"readsInClonesBeforeClustering": 34336,
"readsDroppedWithLowQualityClones": 77,
"clonalChainUsage": {
"type": "chainUsage",
"chimeras": 0,
"total": 25533,
"chains": {
"TRA": {
"total": 15308,
"nonFunctional": 2060,
"isOOF": 2035,
"hasStops": 25
},
"TRB": {
"total": 10224,
"nonFunctional": 635,
"isOOF": 595,
"hasStops": 40
},
"TRD": {
"total": 1,
"nonFunctional": 1,
"isOOF": 1,
"hasStops": 0
}
}
}
}