BCR-Seq of Orictolagus cuniculus: Blood, spleen and bone marrow samples
Double stranded cDNA was synthesized using multiple rabbit immunoglobulin gene specific primers with additional universal sequences and multi-cycle amplification was conducted with universal amplification primers. First strand cDNA was synthesized using 1 g of total RNA and SuperScript  first-strand cDNA synthesis kit with rabbit J gene specific reverse primers with additional restriction or linker sequence, P7 sequence and UMI-barcode (Smith et al., 2017). First-strand cDNA was purified using AmPure XP beads (A63881; Beckman Coulter) at a 1:1.8 ratio, then second-strand cDNA was synthesized using KAPA HiFi HotStart DNA polymerase (KK2502; Kappa Bioscience) with rabbit V gene specific forward primers with additional restriction or linker sequence and P5 sequence (98 for 4 min, 60 for 1 min, 72 for 5 min). Double-stranded cDNA was purified using AMPure XP beads (A63881; Beckman Coulter) at a 1:1.8 ratio, then amplified with KAPA HiFi HotStart DNA polymerase (KK2502; Kappa Bioscience) using two universal primers containing Illumina adapters and index sequences (95 for 3 min, 25 cycles of 95 for 30 s, 65 for 30 s, 72 for 1 min, and 72 for 5 min).
Rabbit_K_Universal