allow "stiching" of regions between others (60 - 150bp)

[jjti]

Tip: “Stitching” Fragments Together using Oligos

When you need intervening sequence between two PCR products, one method is to “stitch” together several oligos. This technique is especially useful for introducing promoters, terminators, and other short sequences into the assembly and is used when the part to be inserted is too long to include on overlapping PCR primers (>60 bp) but too short to make its own part (<150 bp).

Please note that the way to design the “stitching” primers and the amounts of them to include in the Gibson reaction are different than with normal PCR primers. The details are published in ( Nat Methods 2010; 7:901-3 ).