Figure 1

Description of vectors for the Creator and Creator Splice systems. A) Schematic of the donor vector illustrating the essential features of the donor vector. ORFs are cloned into the Multiple Cloning Site (MCS) that is flanked by two loxP sites. The CmR ORF following the MCS, but still within the two loxP sites, ensures that both the ORF cassette from the donor vector and the acceptor vector backbone (containing the CmR promoter) are retained in the final product. The ampicillin resistance gene (AmpR) provides positive selection and the SacB gene imparts negative selection (against the donor backbone). Sequences of the novel MCS for the modified Creator and Creator Splice vectors are indicated below the diagram. The MCS starts at the AscI site and ends at the PacI site. The last four nucleotides of the loxP site are shown for orientation. The splice acceptor sequence in the Creator Splice vector is shown in green to emphasize the difference between the two vectors. B) A schematic of the modular nature of the acceptor vectors. The types of cassettes (promoter, tags, selectable markers) that can be interchanged to create vectors for specific proteomics needs are illustrated, not all combinations are available (Table 2). Abbreviations: AmpR – ampicillin resistance gene, b – bacteria expression, CmR – chloramphenicol resistance ORF, i – insect expression, m – mammalian expression, MCS – multiple cloning site, SA – splice acceptor.