Efficient Rutin and Quercetin Biosynthesis through Flavonoids-Related Gene Expression in Fagopyrum tataricum Gaertn. Hairy Root Cultures with UV-B Irradiation

摘要

Transformed hairy roots had been efficiently induced from the seedlings of Fagopyrum tataricum Gaertn. due to the infection of Agrobacterium rhizogenes . Hairy roots were able to display active elongation with high root branching in 1/2 MS medium without growth regulators. The stable introduction of rol B and aux 1 genes of A. rhizogenes WT strain 15834 into F. tataricum plants was confirmed by PCR analysis. Besides, the absence of vir D gene confirmed hairy root was bacteria-free. After six different media and different sources of concentration were tested, the culturing of TB7 hairy root line in 1/2 MS liquid medium supplemented with 30 g l~(-1)sucrose for 20 days resulted in a maximal biomass accumulation (13.5 g l~(-1)fresh weight, 1.78 g l~(-1)dry weight) and rutin content (0.85 mg g~(-1)). The suspension culture of hairy roots led to a 45-fold biomass increase and a 4.11-fold rutin content increase in comparison with the suspension culture of non-transformed roots. The transformation frequency was enhanced through preculturing for 2 days followed by infection for 20 min. The UV-B stress treatment of hairy roots resulted in a striking increase of rutin and quercetin production. Furthermore, the hairy root lines of TB3, TB7, and TB28 were chosen to study the specific effects of UV-B on flavonoid accumulation and flavonoid biosynthetic gene expression by qRT-PCR. This study has demonstrated that the UV-B radiation was an effective elicitor that dramatically changed in the transcript abundance of ftpAL , FtCHI , FtCHS , FtF3H , and FtFLS-1 in F. tataricum hairy roots.