a8842728-9a41-11e7-9f06-000e1e0af2dc
Zea mays B73, seedling root tip (0-3mm), whole genome sequencing from 2C and 8C sorted nuclei
B73 seedling roots were pulse-labeled with the thymidine analog 5-Ethynyl-2''-deoxyuridine (EdU) for 1 h to allow actively replicating DNA to incorporate the EdU. The terminal 0-3 mm root segments were excised, fixed in 1% formaldehyde for 15 minutes, washed in PBS, and snap frozen. Nuclei were isolated from the frozen, excised roots, the incorporated EdU was "clicked" to Alexa Fluor-488 (AF-488), and total DNA was stained with DAPI. Nuclei were analyzed based on EdU incorporation (AF-488 fluorescence) and DNA content (DAPI fluorescence), and 2C and 8C unlabeled nuclei were sorted. Genomic DNA was extracted from sorted nuclei and Illumina sequenced.