Deposition of diamond inside the trenches or microchannels by chemical vapor deposition (CVD) is limited by the diffusion efficiency of important radical species for diamond growth (H, CH3) and the pore depth of the substrate template. By ultrasonic seeding with nanodiamond suspension, three-dimensional (3D) penetration structure diamond was successfully deposited in cylindrical microchannels of Cu template by hot-filament chemical vapor deposition. Micro-Raman spectroscopy and scanning electron microscopy (SEM) were used to characterize diamond film and the effects of microchannel depth on the morphology, grain size and growth rate of diamond film were comprehensively investigated. The results show that diamond quality and growth rate sharply decrease with the increase of the depth of cylindrical microchannel. Individual diamond grain develops gradually from faceted crystals into micrometer cluster, and finally to ballas-type nanocrystalline one. In order to modify the rapid decrease of diamond quality and growth rate, a new hot filament apparatus with a forced gas flow through Cu microchannels was designed. Furthermore, the growth of diamond film by new apparatus was compared with that without a forced gas flow, and the enhancement mechanism was discussed.