NextSeq 550 paired end sequencing; ATAC-seq of zebrafish primordial germ cells and somatic cells during early embryogenesis
ATAC-seq of zebrafish primordial germ cells and somatic cells during early embryogenesis
Zebrafish PGCs were isolated via FACS after embryo dissociation. The embryos were grown until the desired stage at 28.5C and collected in synchronous batches. Cell dissociation occurred via pipetting thoroughly up and down the pooled embryos until the solution appeared homogeneous. Yolk excess was removed by two rounds of centrifugation at 300 x g (5 minutes). Cells were passed through a 50 Î¼m mesh and loaded into a FACS Aria II machine for cell sorting. Cells were lysed and nuclei extracted as described in Buenrostro et al., 2013. Extracted nuclei were incubate 30 minutes at 37 C in presence of Tn5 transposase (Illumina, FC-121-1030, UK) for tagmentation as described in Buenrostro et al., 2013. Tagmented DNA was purified and amplified by PCR prior to sequencing.  DNA library was size-selected via the Agencourt AMPure XP beads kit (Beckman Coulter, A63880, USA).