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. 2008 Nov 27;3(11):e3827. doi: 10.1371/journal.pone.0003827

Figure 2. Site allocation and compatibilities.

Figure 2

The MCS inserted into the recipient plasmid was assembled from Age I, Spe I, Nhe I, Blp I, Bsr GI, Mlu I, Asc I, Sma I, Pac I, Acl I, and Dra III. PCR donor fragments made with core 2 primers (A) are digested with Mlu I and Asi SI and inserted into Asc I and Pac I (B). Previous insertions (C) are excised with Mlu I and Pac I and sub-cloned into another vector prepared either previously or in parallel with (C) and opened with Asc I and Pac I (D). Entire cloning regions (all cores and inserts) can be shuttled between vectors (E–F) using the external shuttling sites, Age I and Acl I or Dra III. * Blp I and Sma I were included as spacers to distance the two sites to be double digested for receiving inserts.