GSM2575596: scTrioSeq2Met_HEp-2_6; Homo sapiens; Bisulfite-Seq
The single viable cells were individually picked into 200-uL tubes containing lysis buffer. For scTrio-seq2, we used magnetic beads (Invitrogen, cat. #65011) to separate the nucleus and RNA of one single cell. We added 0.2 ?L magnetic beads to each single-cell lysis buffer. Then the single cells were lysed and vortexed for 1 min to release RNA. The lysis products were then centrifuged at 1,000 × g for 5 min at 4°C, and placed on the magnetic rack for 5 min. The magnetic beads can aggregate on the surface of the nucleus to maintain the nucleus in the pellet, while the RNA was released in the supernatants. The supernatants containing RNA were transferred to a new tube for transcriptome sequencing. The remaining beads containing a single nucleus were re-suspended with lysis buffer of scBS-seq for DNA methylation sequencing For single-cell whole-genome bisulfite sequencing, the scBS-seq libraries were constructed according to the published protocol as described in 2017 (Clark et al., 2017). For CRC01 and CRC02, the transcriptome sequencing libraries were constructed according to Tang protocol (Tang et al., 2009); for the remaining patients, the transcriptome sequencing libraries were constructed according to a multiplexed scRNA-seq method, in which the poly T primers were combined with barcodes and unique molecule identifiers (UMIs) (Dong et al., 2018).