Scalable gene synthesis platform schematic for OLS Pool 2Pre-designed oligonucleotides (no distinction is made between dsDNA and ssDNA in the figure) are synthesized on a DNA microchip (a) and then cleaved to make a pool of oligonucleotides (b). Plate-specific primer sequences (yellow or brown) are used to amplify separate Plate Subpools (c) (only two are shown), which contain DNA to assemble different genes (only three are shown for each plate subpool). Assembly specific sequences (shades of blue) are used to amplify assembly subpools (d) that contain only the DNA required to make a single gene. The primer sequences are cleaved (e) using either Type IIS restriction enzymes (resulting in dsDNA) or by DpnII/USER/γ exonuclease processing (producing ssDNA). Construction primers (shown as white and black sites flanking the full assembly) are then used in an assembly PCR reaction to build a gene from each assembly subpool (f). Depending on the downstream application the assembled products are then cloned either before or after an enzymatic error correction step.
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