In this study, the pseudo-trinucleotide composition was proposed, and a novel method was developed by support vector machine (SVM) with the pseudo-trinucleotide composition as input parameter to represent DNA sequence for DNA methylation prediction. The model was evaluated on two datasets, including a dataset of Rollins (dataset_1) and a dataset collected healthy human records from the MethDB database (dataset_2). For dataset_1, the Matthews correlation coefficient (MCC) and accuracy (ACC) by jackknife validation were 0.8051 and 0.6098, respectively. For dataset_2, the MCC and ACC were 0.8500 and 0.7203, respectively. The good prediction results reveal that the pseudo-trinucleotide composition is an effective representation method for DNA sequence and plays a very important role in the prediction of DNA function.