GSM8136393: LT5_LLC-OVA_shNT_IgG_5 (biol rep 5); Mus musculus; OTHER
Total RNA was extracted from the bulk tumor tissues for TCR beta chain library construction using the Direct-zol RNA MiniPrep kit. 1 μg or lower amounts of RNA were converted into cDNA using a cDNA Synthesis Kit (Quantabio) with a constant region-specific primer. A multiplex PCR system was introduced to amplify the CDR3 region of rearranged TCRB loci. A set of forward primers, each specific to one or a set of functional TCR Vb segments, and a reverse primer specific to the constant region of TCRB, were used to generate amplicons covering the entire CDR3 region. PCR products were loaded onto 2.5% agarose gels, and bands centered at 250–400 bp were excised and purified using a QIAquick Gel Extraction kit (QIAGEN).