Fig 1.

Schematic pSpiro vector maps showing constructed cassettes and relevant restriction sites. The episomal vectors contain the ampicillin resistance gene (AmpR) for selection in E. coli and the pac, bsr, and nptII genes for selection in S. salmonicida. The pac and bsr genes are flanked by the 5′ UTR and 3′ UTR of the ornithine carbamoyltransferase (OCT) gene, which are cloned between the HindIII/NcoI and XhoI/KpnI sites, respectively. The nptII gene is inserted between the 5′ UTR and 3′ UTR of the fructose bisphosphate aldolase (FBPA) gene, and the entire cassette is cloned between SpeI and NdeI. The three eukaryotic selection markers can be exchanged using combinations of KpnI/SpeI and NdeI/HindIII. The 3′UTR from the ribosomal protein S15A (RbpS15A) gene was cloned between the ApaI and SacI sites. (A) The C-terminal cassettes (3×HA, 2×OLLAS, 3×MYC, SBP-GST, and SF-TAP) are cloned between the NotI and ApaI sites. Ten unique restriction sites are available for cloning in the MCS region of the pSpiro C-terminal tagging vectors. (B) The N-terminal vectors carry the α-tubulin (αTub) 5′ UTR cloned between the HindIII and EcoRV sites. The N-terminal tagging cassettes (3×HA, 2×OLLAS, SBP-GST, and SF-TAP) are cloned at the EcoRV and BamHI sites. Six restriction sites are available for cloning in the MCS region of the pSpiro N-terminal tagging vectors.