Molecular Cloning and Characterization of Oleoyl-ACP Thioesterase Gene from Brassica Juncea+

Oleoyl-ACP thioesterase (TE) is the chain length-determining enzyme in de novobiosynthesis of oleic acid. For cloning the gene encoding oleoyl-ACP TE from Brassica juncea, a PCR-amplified DMA probe was developed with the primers designed from the known sequences available in the Genbank. It was used to screen a genomic library of B. juncea constructed in λAEMBL 3 to get the complete gene. A 4.0 kb Bam HI fragment from the clone λ5.12 and hybridizing to the probe was sequenced. A stretch of 2606 bp genomic sequence was found to contain an ORF of 1101 bp with ∼ 0.4 kb untranslated regions at both the ends and 5 introns of varied lengths. By homology analysis, the ORF was found to encode an oleoyl-ACP thioesterase gene.The promoter region contains major cis elements required for transcription.