Ekspresi Gen Penyandi Asam o-aminolevulinat Sintase dari Rhodobacter sphaoerides pada Arabidopsis thaliana

Abstract

Synthesis of the tetrapyrrole precursor 8-Aminolevulinate (ALA) in plants starts with glutamate and is a tRNA dependent pathway consisting of three enzymatic steps localized in plastids. In animals, yeast and some of the bacteria, ALA is formed in a single step from succinyl CoA and glycine by aminolevulinate synthase (ALA-S) inside mithochondria. A gene encoding ALA-S from Rhodobacter sphaeroides under 35S promoter was introduced into the genome of Arabidopsis thaliana employing Agrobacterium tumefaciens-mediated transformation in planta. Five putative transgenic lines were obtained and four among them were positive carrying hemA dan kanamycin. All the transgenic lines expressed the transgene at the level of RNA confirmed by RT-PCR analysis. The fresh and dry weigth was increased up to 16.8% and 16.6%, respectively. But, two lines was abnormal, one was reduced in fresh and dry weight because of poor root development (line no 3) and the other reduced in seed production for about one quarter and exhibited late flowering (line no 4). The ALAS activity in line no 4 was 40.5 nmol.mg¹.h¹ and the capacity to synthesize ALA and chlorophyll was increased 153.6% and 43%, respectively. In addition it could also improve tolerance of salinity stress when exposed to 200 mM NaCl. In this study, the gene for Chlorophyll A Oxygenase has been cloned from Arabidopsis thaliana.