Productive assembly has been achieved for DNA fragments with as little as a 12 bp overlap, however, it depends on the GC content of the overlap. We recommend using at least 15 bp overlaps, or more, for dsDNA assembly with a Tm ≥ 48°C (AT pair = 2°C and GC pair = 4°C). Increasing the length of overlap between fragments also reduces the amount of DNA needed for assembly.