GSM6387249: Fus, low, HSCs, tagHi-C, rep1; Mus musculus; Hi-C
Fus low and Fus high HSCs(lineage-, c-kit+ Sca-1+ CD34-) from bone marrow of mice using fus gtp; Young(2 months) and Aged(30 months) HSCs(lineage-, c-kit+ Sca-1+ CD34-) from bone marrow of mice Hi-C: We performed in tagHi-C on FUSlow HSCs and FUShigh HSCs(Cell reports 32, 108206 (2020)). In briefly, 20,000 HSCs were sorted and crosslinked in 100 uL 1% formaldehyde for 10 minutes at room temperature. Cell pellet was collected by centrifuged for 5 minutes at 3,000 g at 4℃ and lysed in 25uL of Hi-C lysis buffer. Next MboI restriction enzyme in 2.5uL of 10x NEB buffer 2 were added to digest chromatin. Then DNA fragments were ligated by incubated in Ligation master mix (66.3 uL of ddH2O, 12 uL of 10 x T4 DNA ligase buffer, 10 uL of 10% (v/v) Triton X-100, 1.2 uL of 100 x BSA and 1 uL of 400 U/uL T4 DNA ligase).Next, DNA was extracted by150 uL Phenol-Chloroform-Isoamyl alcohol (25:24:1). Next, 20 uL of DNA fragmentation mix (4 uL of 5 x TTBL, 5 uL of TTE Mix V1, 5 ng DNA and ddH2O) was used for PCR amplification. After amplification, the samples were purified with AMPure beads once and resolved with ddH2O to prepare for sequencing library after the DNA concentration was quantified with Qubit. The libraries were sequenced using an Illumina Hiseq platform with 150-bp paired-end.