Fig. 6

(a) Drop-test on solid media determining the minimal inhibitory concentration of nourseothricin on C. oleaginosus wt. (b) Growth curves of C. oleaginosus wt with increasing concentrations of nourseothricin, determining the minimal inhibitory concentration in liquid media. (c) Map of the pRF_PPIp_NAT plasmid, which was used for ATMT-derived integration of the NAT resistance gene. Shown elements, besides the described gene cassette, include: KanR, bacterial kanamycin resistance; oriV, bacterial origin of replication; trfA, bacterial plasmid replication initiator protein. Plasmid maps were created with Geneious. (d) Restriped colonies after transformation of the wt control and the transformants on nourseothricin plates. (e) Gel-electrophoresis of NAT amplified by PCR (783 bp) from genomic DNA of the transformants. The gel was cropped for clarity between the marker (M) and the lanes 2 to 5 and after lane 5, the complete gel picture is included in Figure S3. (f) Growth curves of the transformants and the wt, in triplicates, with and without the addition of nourseothricin. NAT, N-acetyl transferase; Nrs, nourseothricin; wt, wild type