Functional Analysis Of GA20-oxidase3Gene In The ABA Signal Transduction Pathway In Arabidopsis

Gibberellin20-oxidase (GA20ox) is the key rate-limiting enzyme in the processof gibberellin (GA) synthesis. GA with activity in plants can break dormancy,promote seed germination, plant growth, flowering and other biological chara-cteristics. GA20ox regulates every stage of plant growth and development by regu-lating gibberellin synthesis. Abscisic acid (ABA) is also important during the deve-lopment of plant. ABA can promote dormacy, inhibit the growth and promote stomatalclosure. Physiological roles of GA and ABA in the plant are just opposite, whichindicates the two ones have antagonistic effects.Therefore, the research of GA20oxtranscript and protein level changes with ABA treatment is very important.In this study, GA20ox3gene in Arabidopsis thaliana were cloned with theIn-Fusion cloning system, luciferase (LUC) as a detection tag was attached at the Nterminal of the target gene. Then this recombinant faction was constructed to theexpression vector pEGAD, and was transformated to Arabidopsis laying the inflo-rescence immersion infection mediated by Agrobacterium. After T1generation ofArabidopsis was screened by Basta and Luminometer, transgenic plants was obtained.Phenotypic analysis found that overpression of GA20ox3is insensitive to ABA,mutant of ga20ox3is sensitive to ABA. Followed by further molecular identification,Western Blot also had a specific band, which indicates that GA20ox3geneoverexpressed successfully in Arabidopsis. For more biological function of this gene,via real-time quantitative PCR, the tissue expression analysis of this gene inArabidopsis plants showed that GA20ox3gene was expressed in various organs, butflowers and roots mostly, which maybe involves with GA specific physiological rolein each organ. In order to study the difference of the levels between transcription andprotein, through GA20ox3under ABA treatment, qPCR result showed the leval of transcription changed, which indicated that the expression of GA20ox3protein waspossiblely regulated by ABA at the leval of transcription.Detection and analysis of various experiments indicated that the expression ofGA20ox3may be regulated by ABA to control the GA synthesis, forming thecorresponding biological traits. The mechanism ABA regulating GA20ox3providesthe theoretical foundation for improving the crop quality.