title: V186.2DJH-Cg1 sequence of NP-CGG immunized mouse fed fiber diet and plain water, sample 3
design: RNA was extracted from mouse spleen cells using the RNeasy Mini Kit (Qiagen). cDNA was synthesized from 1-2 g total RNA with the SuperScript III First-Strand Synthesis System (Invitrogen) using oligo-dT primer. Rearranged V186.2DJH-C1, V186.2DJH-C2b or V186.2DJH-C3 cDNA encoding the anti-NP IgH chain was amplified using a V186.2 leader-specific forward primer together with a reverse C1-, C2b- or C3-specific primer tagged with Illumina clustering adapters and Phusion high fidelity DNA polymerase (New England BioLabs). The amplified library was tagged with barcodes for sample multiplexing, PCR enriched, and annealed to the required Illumina clustering adapters. High-throughput sequencing was performed by Paired-end sequencing (2300 bp) using the Illumina MiSeq platform.