title: GSM7115807: Cis + PBS + aPD-1 replicate 5; Mus musculus; OTHER
library_construction_protocol: Snap-frozen samples of tumors were lysed in Qiazol with a TissueLyser II (Qiagen). RNA was then isolated using the miRNeasy Micro Kit from Qiagen, according to the manufacturer's instructions. Extracted RNA was then amplified using the MessageAmp II aRNA Amplification Kit (Ambion) with the following modifications: in vitro transcription was performed at 37°C for 16h. The resulting cRNA was converted to ssDNA using the Superscript III (Thermofisher) and a collection of TRBV specific primers containing UMI and Illumina adapters. TCRs were then amplified by PCR (20 cycles with the Phusion from NEB) with a single primer pair binding to the constant region and the adapter linked to the TRAV/TRBV primers added during the reverse transcription. A second round of PCR (25 cycles with the Phusion from NEB) was performed to add the Illumina adapters containing the different indexes.