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. 2014 Feb 13;9(2):e88218. doi: 10.1371/journal.pone.0088218

Figure 2. Assembly of Level II, III and IV plasmids.

Figure 2

A) LI modules and LI dummies (LI dy) are fused with compatible BsaI overhangs into a LII backbone vector by BsaI cut-ligation. B) To create a LIII binary vector for plant expression up to five LII synthetic genes (LII 1–2, LII 2–3, LII 3–4, LII 4–5 and LII 5–6) or suitable LII dummies (LII dy) are combined with compatible BpiI overhangs into a LIII backbone by BpiI cut-ligation. C) A LIV plasmid is constructed from LIII multigene assemblies combined with LI dy fragments, which carry compatible BsaI overhangs, into a LII backbone by BsaI cut-ligation. LI fusion sites are named A to G, while LII fusion sites are numbered from 1 to 6. Triangles indicate the orientation of the restriction sites relative to the BsaI and BpiI recognition sites.