CUT&Tag Seq for H3K27me3 of mus musculus: E9.5 embryo neural tissues
Pregnant mice were given corn oil intraperitoneally (0.4 ml) from E6.5 for three consecutive days. On E9.5, embryos were collected and carefully inspected for NTDs under a dissecting microscope. Ten embryos were pooled as a sample for CUT&Tag experiment. Embryonic neural tissues, from the most rostral aspect of the forebrain to the caudal aspect of the hindbrain, were excised. Neural tissues were homogenized and followed by incubation with Concanavalin A-coated Magnetic Beads (Novoprotein, China) for 10 min at room temperature. Next, H3K27me3 rabbit pAb (1:100, Merck Millipore, USA) were employed as primary antibodies, and Goat Anti-Rabbit IgG H&L (1:100, Abcam, USA) was used to bind primary antibodies. After binding pG-Tn5 adapter complex (Novoprotein, China), tagmentation was conducted. Following library amplification, DNA quantity was determined with Qubit 4 (Thermo Fisher Scientific, USA) and library quality characterized with Agilent TapeStation 4200. All libraries were sequenced using NovaSeq 6000 (Illumina, USA).