Fig. 2

DNA hypermethylation results in the low expression of miR-375. a–c The methylation status of miR-375 was analyzed by MSP in 7 leukemic cell lines (a), 40 primary AML blasts (b), and 20 normal controls (c). B: Blank; P: positive control of methylated DNA. Bands of ‘M’ or ‘U’ are PCR products amplified by methylation-specific or unmethylation-specific primers, respectively. Shown are the representative figures for primary AML blasts and normal controls. d A CpG map of the sequenced region was analyzed by MethPrimer software. e Bisulfite genomic sequencing was performed to detect the methylation status of the DNA sequences at − 260 bp − + 136 bp in the pre-miR-375 gene upstream region in HL-60, THP1, and one normal control (NC#1). Five PCR products were shown for each sample. Each row of circles represents the sequence of an individual clone. Black circles and empty circles represent methylated and unmethylated CpG dinucleotides, respectively (Left). Shown was the summary of frequencies of methylated CpG dinucleotides detected in HL-60, THP1, and one NC by bisulfite genomic sequencing (Right). f The expression of miR-375 was detected in HL-60 and THP1 cells treated with 5 μM AZA for 2 and 4 days. *P < 0.01 versus untreated cells