RNA-seq in acute myeloid leukemia (AML) cells with and without knockdown of METTL14

To dissect the mechanism underlying the oncogenic function of METTL14 in AML, we performed deep sequencing for mRNA isolated from MM6 and NB4 cells with and without knockdown of METTL14 We lentivirally transduced pLKO.1-based lentiviral shRNA targeting METTL14 (i.e., shM14-#2) or scramble shRNA (i.e., shNS) into human MM6 and NB4 AML cells. After selected with puromycin (0.5 µg/mL) for two passages, cells were collected and RNA was extracted using TRIzol reagent (Invitrogen, Carlsbad, CA). PolyA RNA was subsequently purified from100 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 duplicate by Illumine Hiseq 1000 with single end 50-bp read length.