GSM4785470: BSPCR_col; Arabidopsis thaliana; OTHER
BS-PCR-seq was performed as previously described (GallegoBartolome et al., 2019). Briefly, leaf tissues from adult plants of representative T2 lines containing the ZF-SssI transgene were collected and DNA was extracted following a CTAB-based method. Bisulfite conversion was done using the Epitect Bisulfite Conversion kit (QIAGEN). The following regions of FWA were analyzed: Region 1 (chr4: 13038143-13038272); Region 2 (chr4: 13038356-13038499); Region3 (chr4: 13038568-13038695), which cover fragments of the promoter and 5′ transcribed region of FWA. Pfu Turbo Cx (Agilent) was used to amplify bisulfite-treated DNA using primers containing Illumina adaptors. The primers used are listed in Table S1. For each sample, individual PCR products from each of the three FWA regions were pooled and purified using AMPure beads (Beckman Coulter) before making the libraries. Libraries were made from purified PCR products using a TruSeq Nano DNA Library Prep kit for Neoprep automated library preparation machine (Illumina), a Kapa DNA hyper kit (Kapa Biosystems) with Illumina TruSeq DNA adapters or an Ovation Ultralow V2 kit (NuGEN). Libraries were sequenced on Illumina HiSeq 2000 or HiSeq 2500.