RNA-seq in murine acute myeloid leukemia (AML) cells with and without knockdown of Tet1

To explore the global targets and dissect the mechanism underlying the oncogenic function of TET1 in AML, we performed deep sequencing for mRNA isolated from mouse MLL-ENL-ERtm cells with and without knockdown of Tet1. We transduced pGFP-V-RS-based retroviral shRNA targeting Tet1 (i.e., shTet1) as reported in Huang H et al., PNAS, 2013 (reported as shTet1-a+b), or scramble shRNA (i.e., shNS), into mouse MLL-ENL-ERtm cells. After puromycin selection (1 µg/mL) for three passages and SILAC adaption, cells were collected and RNA was extracted using miRNeasy kit (Qiagen). PolyA RNA was subsequently purified from 50-100 ng total RNA using NEBNext Poly(A) mRNA Magnetic Isolation Module. NEBNext Ultra Directional RNA Library Prep Kit (New England BioLabs, Ipswich, MA) was used for library preparation. Each group was sequenced in triplicate by Illumina Hiseq 1000 with single end 51-bp read length.