FAQ: What are the shortest overlaps that can be used with this assembly method?

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.
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    Primer Design and Fragment Assembly Using NEBuilder HiFi DNA Assembly or Gibson Assembly®

    Watch an interactive tutorial on primer design to see how simple it really is to clone with either NEBuilder HiFi DNA Assembly or the Gibson Assembly Cloning Kit.