GSM7956888: H3 ChIP seq HSVd 20 dpi bioreplicate 2; Cucumis sativus; ChIP-Seq
Immunoprecipitated DNA was extracted using the GeneJET PCR Purification Kit (Thermo Fisher), following the manufacturer's instructions. 500 mg of systemic leaves were chemically cross-linked using 1% formaldehyde. Then, nuclei were isolated from cross-linked material following a standard nuclei isolation protocol based on sucrose gradients as previously described (Moreno-Romero et al., 2016). The resuspended nuclei pellets were sonicated for 9 cycles of 20 s On and 45 s Off at 4 °C and high power to obtain the chromatin. Afterward, the Immunoprecipitation (IP) was performed following a standard IP protocol and using the following antibodies: H3 (Reference: 07-690, Merck), H3K9me2 (Reference: pAb-060-050, Diagenode) and H3K27me3 (Reference: 07-449, Merck). DNA libraries were prepared using the NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® (New England Biolabs) and each individual library was barcoded using the NEBNext® Multiplex Oligos for Illumina® kit (New England Biolabs).