GSM4648107: K3; Mus musculus; RNA-Seq
RNA and DNA methylation were prepared for sequencing RNA was isolated from mII and amplified using Smart-Seq v4 Ultra Low Input RNA Kit (Clontech Laboratories,Mountain View, CA, USA) and sequenced by Novogene Corporation (Haidian, Beijing, China). DNA was extracted using the oral swab gene extraction kit TIANamp Genomic DNA Kit (TianGen); the egg cell sample had fewer cells and was directly lysed with proteinase K (Qiagen). The genome DNA or oocytes' lysate is digested with MspI / XmaI (Fermentas) double digestion. At the same time, Unmethylated λDNA of about 1% of the sample DNA mass should be added to the digestion system as an internal reference to investigate the conversion rate. Digestion products are used for terminal repair, addition of dA, and methylated linker connection (NEB, all cytosines in the linker sequence are methylated); the ligation product is converted to bisulfite and the conversion product is purified and recovered, referring to EZ DNA Methlyation -Direct Kit (ZYMO) instructions; the transformation products are amplified by 2X Kapa HiFi U + master Mix (Kapa), Universal primer (NEB), Index primer (NEB); the products are screened by XP magnetic beads (Beckman Coulter) 200- The 450bp fragment was finally sequenced with HiSeq X10 analyzer (Illumina) 150 bp-read pair-end (PE).
GEO Accession
GSM4648107