Li, C.; Saimaiti, Y.; Li, M.; Wang, S.; Yin, F.; Fang, Z.; Jia, S.; Wang, F.; Ye, D.; Wang, L.; Zuo, X.; Fan, C.

High-throughput DNA synthesis has revolutionized synthetic biology, molecular diagnostics, genome engineering, and DNA data storage by enabling the scalable and precise construction of nucleotide-based information systems. However, existing high-throughput synthesis technologies rely on top-down fabrication strategies to maximize throughput, yet they are inherently constrained by physical limitations. These constraints make it exceedingly difficult to achieve single-molecule level synthesis control that is essential for advancing next-generation DNA synthesis. Here, we present a DNA framework-based bottom-up enzymatic synthesis strategy that enables single-molecule level control of DNA synthesis with high-throughput. Leveraging the nanoscale addressability of DNA frameworks, we achieved a synthesis site pitch of 10.9 nm, 183-fold smaller than that of previously reported electrode arrays. Theoretical projections suggest that this approach could scale synthesis throughput to 530 billion sequences per square centimeter, representing a four-order-of-magnitude improvement over existing synthesis arrays. This approach lays the foundation for gigabyte-scale DNA writing and offers a highly accessible pathway toward large-scale DNA synthesis for emerging applications such as molecular data storage.