FIG 3.

Evaluation of autonomously replicating activity and plasmid retention in P. pastoris. (A) EGFP expression plasmids containing Cen2 (pUC19-Cen2-EGFP), LOR2-CC2 (pUC19-LOR2CC2-EGFP), LOR2 (pUC19-LOR2-EGFP), LOR2(1–111) [pUC19-LOR2(1–111)-EGFP], or PARS1 (pUC19-PARS1-EGFP). (B) CBS7435 harboring pUC19-Cen2-EGFP (Cen2), pUC19-LOR2CC2-EGFP (LOR2-CC2), pUC19-LOR2-EGFP (LOR2), pUC19-LOR2(1–111)-EGFP [LOR2(1–111)], and pUC19-PARS1-EGFP (PARS1) was grown in BMGY medium with or without 200 μg/ml Zeocin for 24 h. The strains CBS7435 (control) and T38473-EGFP (genomic integration) were grown in BMGY medium without Zeocin for 24 h. The GFP expression level of each cell was measured by FCM, and the histogram data from 10,000 cells per strain are shown. For each strain, three different transformants were analyzed, and one typical histogram is displayed.