RESUMO
To investigate the effects of the expression of coumaroylquinate 3’-monooxygenase (C3’H) and shikimate O-hydroxycinnamoyltransferase (HCT) in the chlorogenic acid-producing pathway and active ingredients in Chrysanthemum morifolium under flooding stress, we cloned two C3’H genes which were CmC3’H1 and CmC3’H2 and two HCT genes which were CmHCT1 and CmHCT2 by the RT-PCR from Hangju and conducted bioinformatics analysis. During the flower bud differentiation stage, we flooded the C. morifolium and then used β-actin as the reference gene to detect the relative expression of the four genes by the qRT-PCR. Finally, the content of downstream products and other indicators of these four genes in C. morifolium were measured by HPLC. We obtained the four genes’ complete open reading frame and predicted the relative molecular mass of the amino acid sequence and the theoretical isoelectric point (pI). And the protein tertiary structure models were constructed. The qRT-PCR results showed that flooding the C. morifolium for 3 days during the flower bud differentiation stage resulted in significant expression changes of the four genes at different growth stages. The results of HPLC showed that chlorogenic acid, the downstream product catalyzed by the C3’H and the HCT, was significantly higher than that in the control group. It was also found that the content of luteoloside and 3,5-O-di-caffeoylquinic acid was also significantly higher than those of the control group. Therefore, C. morifolium regulates the synthesis of downstream products by regulating the expression of the four genes under flooding stress, thereby responding to flooding stress. And the flooding stress during flower bud differentiation can significantly enhance the accumulation of active ingredients of C. morifolium.
RESUMO
To investigate the effects of the expression of flavonoid 3' hydroxylase gene ( and active ingredients in under flooding stress, we cloned F3'H from Hangju (temporarily named ) and conducted bioinformatics analysis. During the flower bud differentiation stage, we flooded the and then used the Real-time PCR to detect the relative expression of ; Finally, active ingredients of the inflorescence were measured by HPLC.The sequencing results showed that 1 562 bp sequence was acquired with the largest open reading frame of 1 527 bp, which encoded 508 amino acids. The phylogenetic tree found that was highly homologous to other species of Compositae. Real-time PCR results showed that had a significant response to flooding stress and had the highest expression level after flooding for 24 h, which was about 9 times as that of the control group. The results of HPLC showed that luteolin and luteoloside, the downstream products catalyzed by the F3'H, were significantly higher than those in the control group. It was also found that the contents of chlorogenic acid and 3,5- acid were also significantly higher than those of the control group. Therefore, regulates the synthesis of downstream products by regulating the expression of in the flavonoid synthesis pathway under flooding stress, thereby responding to flooding stress. The flooding stress during flower bud differentiation can significantly enhance the accumulation of active ingredients.