MOLECULAR CLONING AND TRANSCRIPT PROFILING OF ASCORBATE OXIDASE GENE AT DIFFERENT GROWTH DEVELOPMENTAL STAGES FROM THERAPEUTICALLY IMPORTANT PLANT SEABUCKTHORN HIPPOPHAE RHAMNOIDES L.

摘要

Ascorbic acid (vitamin C) is a well-known molecule for its nutritional importance. However, the major aspects of its metabolic processes as well as a few of its functions in plants are poorly understood. A candidate gene AO ( Ascorbate oxidase ) known to be involved in ascorbate biosynthesis and metabolism was therefore chosen in seabuckthorn ( Hippophae rhamnoides L.). In this study, full length cDNA sequence of Hr-AO was amplified and cloned through RT-PCR. The amino acid residues encoded by Hr-AO (2160 bp) were 719aa. There was a difference in length of newly isolated cDNA sequence as compared to tomato cDNA with 87% gene homology. Gateway cloning technique was also used to transfer Hr-AO gene to expression vector for functional study. Expression analysis of this gene sequence from six different tissues including vegetative bud, seed, shoot apex, green leaves, green fruit and mature (orange red) fruits showed maximum transcript accumulation in green leaf and young green fruit tissues. This is the first report on description of relationship between expression of Hr-AO and fruit development in such type of bush plant. This new gene isolated from seabuckthorn will help to understand the regulatory role of this enzyme in ascorbic acid metabolism. The investigation suggested that this gene could merely contribute toward ascorbic acid function or may be specific for further genetic engineering of crops.