Cloning And Function Analysis Of GPAT6Gene From Brassica Napus

sn-Glycerol-3-phosphate acyltransferase is a key enzyme that catalyzes the initial step of TAG biosynthetic pathway, and is involved in many processes including plant growth, development and response to abiotic stresses. This study focus on the following4aspects of BnGPAT6gene.1. Two B. napus GPAT6homologs were cloned from leafs of the cultivar xiangyou15using RT-PCR, and were designated as BnGPAT6-1and BnGPAT6-2, respectively. The coding DNA sequences (CDS) of two BnGPAT6genes are1506bp in length, encoding two different polypeptides of501amino acid residues. The proteins were predicted to consist of a haloacid dehalogenase-like hydrolase domain and a lysophospholipid acyltransferase domain. Multiple sequence alignments and phylogenetic analysis of GPAT proteins showed that BnGPAT6-1and BnGPAT6-2have high similarity with GPAT6genes from B. rapa, B. oleracea, Arabidopsis thaliana and A. lyrata.2. Tissue expression amounts of BnGPAT6genes showed that their mRNA are more abundant in flower than other organs, and the patterns rise up at first and then down in the developing embryo. The expression of BnGPAT6genes are inhibited by ABA, while go up simultaneously under drought and6-BA conditions. In the treatment of salt, the expression of BnGPAT6genes are uptrend in a short time and then down, and there isn’t obvious change under stress of water logging.3.Both of the two BnGPAT6genes are constructed into yeast expression vector pYES2.0, and transformed into yeast line INVSC1. Gas chromatography analysis indicates that C14:0fatty acid of both of the BnGPAT6-1and BnGPAT6-2transgenic lines are twice as the abundance of the yeast transformed by YES2.0vector only. However, the content of others fatty acid such as C16:0, C16:1, C18:0and C18:1turn down obviously.4. BnGPAT6-2gene is placed under the control of Napin promoter which is a seed-special promoter, and is transformed into agrobacterium. The special vectors are transformed into wild type Arabidopsis, and the fatty acid are detected by Gas chromatography. The result of Gas chromatography show that the fatty acid component of Arabidopsis seeds are changed by the enzyme coded by BnGPAT6gene.