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The natural yield of cis-1,4-polyisoprene rubber from Hevea brasiliensis is complex and is affected by the tapping-cut height, i.e., the height at which the laticifer site is severed. From rubber biosynthesis to latex exploitation, genetic modifications have been used to improve yield levels. In this study, we determined the correlation between DNA methylation and the fluctuation in rubber yield and its related characteristics during latex regeneration, and cloned and sequenced methylated sites detected by methylation-sensitive amplified polymorphisms. Latex yield and absolute and relative dry yields were lower from high tapping-cut than low tapping-cut plants; latex DNA methylation levels were greater in high tapping-cut than in low tapping-cut plants. There were significant positive correlations between tapping-cut height and latex yield, absolute and relative dry rubber yields, and the hemi-, full, and total methylation ratios in latex. There were significant negative correlations between tapping-cut height and full and total methylation ratios in leaves. Latex yield and absolute and relative dry rubber yields were negatively correlated with full and total methylation ratios in latex, and positively correlated with hemi- and total methylation ratios in leaves. A homology analysis of the methylated genes revealed that the functions of their homologous sequences were mainly associated with transcription, protein biosynthesis, signal transduction, energy, transport, and plant defense. Thus, the accumulation of polyisoprene in the laticiferous production system of Hevea might be regulated by DNA methylation. This is the first report on latex and leaf global DNA methylation analyses during rubber production in self-rooting clones, and thus it provides novel insights into increasing the production of Hevea rubber.