Figure 6. Expression and Lipid analyses of CpFAH- and MALCE1-co-expression lines.

(a) Structure of MALCE1-expression plasmid. MALCE1 gene was fused with a chloroplast transit signal from the CgpsbO gene and cloned under the control of the NO₃⁻-inductive CgNR promoter. A HindIII site for linearisation and primer sites (arrow heads) using genomic PCR are shown. (b) Expression level of exogenic MALCE1 and endogenic CgNR normalised by expression of the endogenous α-tubulin gene in a transgenic line, Cp4-ML47 isolated from the second transformation of the MALCE1-expression plasmid to Cp4 line. The cells were cultured in normal Daigo’s IMK medium containing NO₃⁻ (filled circles) or modified Daigo’s IMK medium containing NH₄⁺ (open circles). (c) Change in ricinoleic acid (RA) amount in the Cp4-ML47 and parental Cp4 lines. P_CgNR, promoter of Chaetoceros gracilis nitrate reductase (CgNR) gene; P_CgACAT, promoter of C. gracilis acetyl-CoA acyltransferase gene; CgpsbO-tp, chloroplast transit signal of CgpsbO gene; Sh ble, Zeocin-resistance gene.