GSM2790120: WT PerC B-1a Rep1; Mus musculus; OTHER
Cells were sorted from 6-8 week old mice. Genomic DNA was extracted from lysed cells and this gDNA was used as template for PCR prior to library prep/adaptor ligation. Primers were used to amplify sucessfully rearranged BCR heavy chains and size selected by gel extraction for 120-280bp from an agarose gel. PCR products were end-repaired using a combination of T4 PNK and T4 polymerase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of preannelaed Illumina compatible adapters, NEXTflex DNA barcodes (Bioo Scientific) which have a single 'T' base overhang at the 3’ end. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on an Illumina HISeq 4000 following the manufacturer's protocols.