GSM6617401: VH1-2_VK1-33-CS_TdT_SP_IgK_4; Mus musculus; OTHER
Splenic B cells used for HTGTS-rep-seq were purified from unimmunized 5-8 weeks old mice by MACS® Microbeads according to the manufacturer's protocol. In brief, spleens were dissected out from unimmunized mice, prepared into single cell suspensions and stained with anti-B220 Microbeads for 20 minutes at 4°C. The splenic B cells were collected using the LS column and MACSTM Separater. GC B cells used for LAM-HTGTS-Rep-SHM-Seq were purified from 8-12 weeks old mice after eOD-GT8 60mer immunization. GC B cells were sorted for the phenotype B220+ (BV711: BioLegend 103255), CD95+ (PE-Cy7: eBioscience 557653) and GL7+ (PE: BioLegend 144607). Genomic DNA from sorted cells was prepared using a DNeasy Blood and Tissue Kit (Qiagen) according to the manufacturer's protocol. The libraries of LAM-HTGTS-Rep-SHM-Seq was constructed as previous described (Chen et al., 2020) and the libraries of HTGTS-Rep-seq was constructed as previous described (Lin et al., PNAS 2016)