GSM4057598: BS-seq replicate1 of LR stage 0; Oryza sativa Japonica Group; Bisulfite-Seq
Twelve µm sections of crown, lateral, and embryonic root initials tissues were put onto RNase-free PEN membrane glass slide (Carl Zeiss, 415190-9041-000). Lasser catpture microdissection (LCM) was performed using the ZEISS PALM MicroBeam system.The RNA of dissected tissues was isolated with RNeasy Plus Micro Kit (Qiagen, 74034). The DNA of dissected tissues was isolated with DNA Micro Kit (Qiagen, 56304). For RNA-seq library construction, Smart-seq v4 Ultra Low Input RNA Kit (Takara, 091817) was used for amplifying purified mRNA. The amplified DNA was sheared to 200-500 bp range by sonication and purified with AMPure XP beads. 1 μg DNA was used for following library construction. Purified DNA was end repaired, added with adenylate in the 3' ends, ligated with adapters, and amplified with PCR for 7-8 cycles with the reagents from TruSeq® ChIP Sample Preparation Kit (Illumina). For BS-seq library construction, Bisulfite conversion was performed by utilizing Zymo EZ Methylation Direct Kit (Zymo, D5020). Converted DNA was purified with PureLink PCR Purification Column Kit (Invitrogen, K210012) with minor modification (add M-Desulphonation Buffer to the column and hold for 15 mins before washing) as Clark et al. (2017) suggested. The converted ssDNA was then used for following BS-seq libraries construction with TruSeq® DNA Methylation Kit (Illumina).
GEO Accession
GSM4057598