• Korean Journal of Plant Tissue Culture
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• v.24 no.2
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• pp.93-97
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• 1997

For the molecular genetic study of cold tolerance mechanism in plants, a cDNA encoding fatty acid desaturase (fad3), converting linoleic acid (18:2, $\omega$-6) to linolenic acid (18:3, $\omega$-3), was isolated from $\lambda$ZAPII Arabidopsis thaliana cDNA expression library by plaque hybridization using fad3 cDNA probe derived from Brassica napus. A 1.8 kb-EcoRI fragment from a lambda clone showing a strong positive hybridization signal was subcloned into pGEM7 and analyzed for its nucleotide sequence. From deduced amino acid sequences, the fad3 gene was revealed to have an open reading frame(ORF) consisting of 386 amino acids with a molecular mass of 44,075 Da. The fad3 gene was compared to chloroplast $\omega$-3 fatty acid desaturase (fad7) and endoplasmic reticulum Δ12 fatty acid desaturase (fad2) to show 70% and 58% amino acid sequence homology, respectively, Especially, amino acids of internal (82 to 151) and carboxy terminal (276 to 333) regions were highly conserved, implying their requisite role for enzymatic functioning of fatty acid desaturases. IPTG-induced fad3 cDNA expression in E. coli cells was suggested to be toxic to bacterial growth.

• Journal of Plant Biotechnology
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• v.1 no.1
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• pp.20-30
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• 1999

In order to modify lipid production of Perilla qualitatively as well as quantitatively by genetic engineering, genes involved in carbon metabolism were isolated and characterized. These include acyl-ACP thioesterases from Perilla frutescens and Iris sp., four different $\beta$-ketoacyl- ACP synthases from Perilla frutescens, and two $\Delta$15 a-cyl-ACP desaturases(Pffad7, pffad3). Δ15 acyl-ACP desa turase (Δ15-DES) is responsible for the conversion of linoleic acid (18:2) to $\alpha$-linolenic acid (ALA, 18:3). pffad 3 encodes Δ15 acyl-desaturase which is localized in ER membrane. On the other hand, Pffad7 encodes a 50 kD plastid protein (438 residues), which showed highest sequence similarity to Sesamum indicum fad7 protein. Northern blot analysis revealed that the Pffad7 is highly expressed in leaves but not in roots and seeds. And Pffad3 is expressed throughout the seed developmental stage except very early and fully mature stage. We constructed Pffad7 gene under 355 promoter and Pffad3 gene under seed specific vicillin promoter. Using Pffad7 construct, Perilla, an oil seed crop in Korea, was transformed by Agrobacterium leaf disc method. $\alpha$-linolenic acid contents increased in leaves but decreased in seeds of transgenic Perilla. Currently, we are transforming Perilla with Pffad3 construct to change Perilla seed oil composition. We isolated three ADP-glucose pyrophosphorylase (AGP) genes from Perilla immature seed specific cDNA library. Nucleotide sequence analysis showed that two of three AGP (Psagpl, Psagp2) genes encode AGP small subunit polypeptides and the remaining (Plagp) encodes an AGP large subunit. PSAGPs, AGP small subunit peptide, form active heterotetramers with potato AGP large subunit in E. coli expressing plant AGP genes.