Molecular Cloning And Functional Analysis Of Flavonoids Biosynthesis Related Genes From Rosa Rugosa

Rose(Rosa rugosa) has an important economic value with brightly colored flowers and fragrance overflows person. The petal of rose is major material sources for extract perfume, the high quality of rose essential oils is expensive, so it is often called “liquid gold”. The rose also has the pharmacological effects of antioxidation and anti-inflammatory, etc. However, the research on rose is mostly limited to cultivation physiology and selective breeding. With advances in rose genetic transformation, it is possible to improve product added value of rose by biotechnology. The research topic is to study the molecular mechanism of flavonoids synthesis, using rose as the main materials. We also isolated Rr DFR1, Rr FLS1 and Rr ANR from rose’s petal and did further study on their function. Major findings are as follows:1 By analysis of the pigment content of roses Rosa rugosa and Rosa multiflora, peach(Prunus persica), carnation(Dianthus caryophyllus), azalea(Rhododendron simsii), camellia(Camellia japonica) and petunia(Petunia hybrida) with red and white flowers, it is found that flavonols could be detected in red and white flowers, but anthocyanins were almost undetectable in the white cultivar. Here, we found that opposing expression of flavonol synthase(FLS) and dihydroflavonol-4-reductase(DFR) genes determined the accumulation of flavonols and anothcyanins in red and white flower of seven species from varied phylogenetic location in plants, and provided a common mechanism for determining flower pigmentation. In which, a series of FLS and DFR genes were isolated from roses, peach, carnation, azalea, camellia and petunia respectively. Expression of many of those FLS genes in tobacco demonstrated the conservation of function, with promoting flavonol biosynthesis and inhibiting anthocyanin accumulation, and resulting in white flowers. Conversely, expression of those DFR genes in tobacco displayed down-regulation of the endogenous Nt FLS gene, and the promotion of anthocyanin synthesis. We demonstrates a correlation between anthocyanin accumulation and the expression ratio of FLS to DFR genes, and the antagonized expression between FLS and DFR genes directs the biosynthesis of flavonols and anthocyanins, thereby determining white versus red coloration of flowers in varied plants species.2 We also isolated Rr ANR from rose by FPNI-PCR to construct plant expression vector, then translated it to tobacco. Over-expressing Rr ANR in tobaccos resulted in pure white flower, an increased accumulation of both PAs and abscisic acid(ABA), and enhanced stress tolerance as well. The transcriptome analyses of the Rr ANR transgenic tobacco and control indicated that a total of 1586 candidate genes were up-regulated in the Rr ANR transgenic tobacco, including genes related to stress reponse and secondary metabolite synthesis. It should be noted that ABA pathway and antioxidant system were activated in the transgenic tobacco. To reveal the molecular mechanism of the enhanced tolerance of Rr ANR transgenic tobacco, Nt ABF from tobacco was isolated. Overexpression of Nt ABF in tobacco up-regulated ROS–scavenging genes and increasd the oxidative tolerance. We also found that the up-regulated ROS–scavenging genes and Nt ABF in PAs- and ABA-treated tobacco. Taken together, overexpression of Rr ANR results in plant tolerance to oxidative stress via scavenging ROS and modulating ABA signaling in plants.