This paper documents the engineering of Arabidopsis thaliana for the ectopic over-expression of SrKA13H (ent-kaurenoic acid-13 hydroxylase)
cDNA from Stevia rebaudiana. HPLC analysis revealed the significant accumulation of
steviol (1-3 μg g(-1) DW) in two independent transgenic Arabidopsis lines over-expressing SrKA13H compared with the control. Independent of the
steviol concentrations detected, both transgenic lines showed similar reductions in endogenous bioactive
gibberellins (GA1 and GA4). They possessed phenotypic similarity to
gibberellin-deficient mutants. The reduction in endogenous
gibberellin content was found to be responsible for
dwarfism in the transgenics. The exogenous application of GA3 could rescue the transgenics from
dwarfism. The hypocotyl, rosette area, and stem length were all considerably reduced in the transgenics. A noteworthy decrease in pollen viability was noticed and, similarly, a retardation of 60-80% in pollen germination rate was observed. The exogenous application of
steviol (0.2, 0.5, and 1.0 μg ml(-1)) did not influence pollen germination efficiency. This has suggested that in planta formation of
steviol was not responsible for the observed changes in transgenic Arabidopsis. Further, the seed yield of the transgenics was reduced by 24-48%. Hence, this study reports for the first time that over-expression of SrKA13H
cDNA in Arabidopsis has diverted the
gibberellin biosynthetic route towards
steviol biosynthesis. The Arabidopsis transgenics showed a significant reduction in endogenous
gibberellins that might be responsible for the
dwarfism, and the abnormal behaviour of pollen germination and seed set.