The invention provides methods for rapid fabrication and use of replicable, high-density nucleic acid arrays. Array features are produced through amplification on a support matrix of nucleic acid templates of interest. A significant advance over conventional DNA array technologies, the so-called polymerase colony ("polony") technology requires no prior characterization or isolation of sequences to be placed on an array, and so is ideally suited to the analysis of unfractionated nucleic acid pools, such as from patient samples or other biological materials. Array-based nucleic acid sequencing methods and reagents are provided. The invention is useful as a genomics research tool (e.g., for whole genome or pedigree analyses; tissue, temporal or cross-species gene expression studies) and in clinical diagnostic, food and environmental testing applications.