DNA is an emerging storage medium for digital data but its adoption is hampered by limitations of phosphoramidite chemistry, which was developed for single-base accuracy required for biological functionality. Here, we establish a de novo enzymatic DNA synthesis strategy designed from the bottom-up for information storage. We harness a template-independent DNA polymerase for controlled synthesis of sequences with user-defined information content. We demonstrate retrieval of 144-bits, including addressing, from perfectly synthesized DNA strands using batch-processed Illumina and real-time Oxford Nanopore sequencing. We then develop a codec for data retrieval from populations of diverse but imperfectly synthesized DNA strands, each with a ~30% error tolerance. With this codec, we experimentally validate a kilobyte-scale design which stores 1 bit per nucleotide. Simulations of the codec support reliable and robust storage of information for large-scale systems. This work paves the way for alternative synthesis and sequencing strategies to advance information storage in DNA.
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