DNA fragmentation is an essential process in developing genetic sequencing strategies, genetic research, as well as for the diagnosis of diseases with a genetic signature like cancer. Efficient on-chip DNA fragmentation protocols would be beneficial to process integration and open new opportunities for microfluidics in genetic applications. Here we present an acoustic microfluidic chip comprising an array of ultrasound-actuated microbubbles located at dedicated positions adjacent to a channel containing the DNA sample solution. The efficiency of the on-chip DNA fragmentation process arises mainly from tensile forces generated by acoustic streaming near the oscillating bubble interfaces, as well as a synergistic effect of streaming stress and ultrasonic cavitation. Acoustic microstreaming and the pressure distribution in the DNA channel were assessed by finite element simulation. We characterized the bubble-enhanced effect by measuring gene fragment size distributions with respect to different ultrasound parameters. For optimized on-chip conditions, purified lambda (lambda) DNA (48.5 kbp) could be disrupted to fragments with an average size of 2 kbp after 30 s and down to 300 bp after 90 s. Mouse genomic DNA (1.4 kbp) fragmentation size decreased to 500 bp in 30 s and reduced further to 250 bp in 90 s. Bubble-induced fragmentation was more than 3 times faster than without bubbles. On-chip performance and process yield were found to be comparable to a sophisticated high-end commercial system. In this view, our new bubble-enhanced microfluidic approach is a promising tool for current and next generation sequencing platforms with high efficiency and good capacity. Moreover, the availability of an efficient on-chip DNA fragmentation process opens perspectives for implementing full molecular protocols on a single microfluidic platform.