GSM8387384: kras_s17n_dna_barcode_GCTATAAGCCCCCTACAAAG; Mesocricetus auratus; OTHER
The cells collected during the directed evolution experiments were harvested, and total RNA was extracted using TRIzol™ Reagent (Thermo Fisher Scientific, #15596018). Subsequently, approximately 3 μg of total RNA was reverse transcribed using the HiScript III 1st Strand cDNA Synthesis Kit (Vazyme, #R312-02) with a primer (SINDBIS_GSPR2: TGCGCCTCCTCGGAAATACG) located in the 3′ untranslated region of the replicative RNA. The KRAS (S17N)-T2A-PuroR barcode region was amplified from both the DNA library plasmid and the cDNA libraries using indexed primers. The purified PCR products were mixed and subjected to adapter ligation, fragment selection, primer and polymerase binding, followed by sequencing using a PacBio Sequel IIe instrument for Circular Consensus Sequencing (CCS) reads acquisition.