4C-seq of Zea mays Pl1-Rhoades (Pl-Rh) seedling rep 1
Library strategy: 4C. Interacting chromatin was isolated from husk and seedling tissues as described for 3C experiments (Hvel et al., 2012, 58: 204 Methods; Louwers et al., 2009, 21: 832 Plant Cell; Weber et al., 2017, 1675: 247 Methods in Molecular Biology). In short, 2-3g tissue was crosslinked, digested with DpnII, de-crosslinked, digested with 50 U NlaIII, re-ligated and amplified using genomic primers fused to P5 (forward) and P7 (reverse) illumina sequencing adapters; a forward primer immediately upstream of the Pl1-Rhoades CDS, and a reverse primer at the 5 end of the doppia transposon fragment upstream of Pl1-Rhoades facing away from the gene. Proper 4C amplicons contained: P5 - forward primer (bait sequence) - DpnII - interacting sequence - NlaIII - reverse primer - P7. For detailed library prep info, see manuscript methods. FP: AATGATACGGCGACCACCGAACACTCTTTCCCTACACGACGCTCTTCCGATCTgattttaaatacgccggcggagctcgatc RP: CAAGCAGAAGACGGCATACGAgcacctattaataattcacagcactgacg