Isolation and partial characterization of beta ketoacyl-acp synthase I (kas I) cDna clone from Chlorella sp.

Abstract

Beta ketoacyl-ACP synthase I (KAS I) is a fatty acid synthase (FAS) that catalyze the elongation of growing fatty acid chains from butyryl-ACP (C4:0) to palmitoyl-ACP (Cl 6:0). In this study, the partial-length of KAS I cDNA clone was isolated and characterized to understand the regulatory mechanism of fatty acid biosynthesis pathway in Chiarella sp. at molecular level. Four heterologous forward primers designed from the conserved regions of the KAS I gene were used in combination with KPN-adaptor reverse primer to amplify the corresponding 3'-end region of the gene by using RT-PCR technique. PCR amplification successfully produced five putative DNA fragments with size ranging between 600 bp to 1000 bp. Cloning and sequencing of the fragments revealed that clone designated as pKASI-Ch5 showed significant (74-75%) nucleotide and amino acid sequence identities to KAS I gene from various plant species such as Helianthus annus, Glycine max and Arabidopsis thaliana. The pKAS1-Ch5 clone (1008 bp) consists of 639 bp open reading frame (ORF) encoding a partial KAS I polypeptide of 213 amino acids with a 369 bp long 3 '-untranslated region. This sequence had been registered in GenBank under accession number EU590913. The isolation of this KAS I cDNA clone is a significant step towards the genetic manipulation of marine microalgae to enhance the production of saturated fatty acids for biodiesel industry.