Cas9-targeted Nanopore sequencing and DNA methylation profiling of full-length glutenin genes in a big cereal genome
nCATS library preparation for Nanopore sequencing was carried out according to the previously published protocols (Gilpatrick et al. 2020; Kirov et al. 2021) and using SQK-LSK109 (Oxford Nanopore Technologies). Briefly, RNP assembly was carried out using 200 ng of each sgRNA and 8 pmol Cas9 protein (Biolabmix, Russia). ~3 g of genomic DNA was used per library. The samples were purified by 0.3 volume of AMPure XP Beads (Beckman Coulter, cat-alog no. A63881) and washed twice by SFB buffer (Oxford Nanopore Technologies, catalog no. SQK-LSK109).