Characterization of DATE in the HGF gene promoter region. (A) and (B) Representative DNA sequencing traces of patients MM 14 and MM 22 with DATEs of 29 and 15 nucleotides, respectively. DATEs of individual patients were amplified by nested PCR, cloned into TA cloning vector pCR2.1 and sequenced using M13 standard primers. (C) HGF mRNA levels in CD138+ cells are plotted against the number of nucleotides present in DATE of the corresponding samples. CD138+ cells purified from bone marrow of myeloma patients were used to quantify HGF mRNA levels by real-time PCR. Corresponding samples were used to isolate genomic DNA for sequencing of DATE in the HGF promoter region. Data shown are HGF mRNA mean fold change ± standard deviation and the number of nucleotides quantified by sequencing.