Establishment of Hairy Root Cultures by Agrobacterium Rhizogenes Mediated Transformation of Isatis Tinctoria L. for the Efficient Production of Flavonoids and Evaluation of Antioxidant Activities

Abstract

In this work, Isatis tinctoria hairy root cultures (ITHRCs) were established as an alternative source for flavonoids (FL) production. I. tinctoria hairy root line V was found to be the most efficient line and was further confirmed by the PCR amplification of rolB, rolC and aux1 genes. Culture parameters of ITHRCs were optimized by Box-Behnken design (BBD), and eight bioactive FL constituents (rutin, neohesperidin, buddleoside, liquiritigenin, quercetin, isorhamnetin, kaempferol and isoliquiritigenin) were quali-quantitatively determined by LC-MS/MS. Under optimal conditions, the total FL accumulation of ITHRCs (24 day-old) achieved was 438.10 μg/g dry weight (DW), which exhibited significant superiority as against that of 2 year-old field grown roots (341.73 μg/g DW). Additionally, in vitro antioxidant assays demonstrated that ITHRCs extracts exhibited better antioxidant activities with lower IC50 values (0.41 and 0.39, mg/mL) as compared to those of field grown roots (0.56 and 0.48, mg/mL). To the best of our knowledge, this is the first report describing FL production and antioxidant activities from ITHRCs.

Induction of hairy roots by Agrobacterium rhizogenes -mediated transformation of spine gourd ( Momordica dioica Roxb. ex. willd) for the assessment of phenolic compounds and biological activities

“…Consistently, hairy roots were exhibited of higher antioxidant activity in D. moldavica (Weremczuk-Jezyna et al, 2013), P. multiflorum (Thiruvengadam et al, 2014a), M. charantia (Thiruvengadam et al, 2014b), and Isatis tinctoria (Gai et al, 2015). Means ± standard deviation of three replicates (n = 3) within a column a-d , or row x-z followed by the same letters are not significantly different according Duncan's multiple range test at P ≤ 0.05.…”

“…Similarly, hairy root cultures of M. charantia have also been cultured in 3% sucrose supplemented MS medium (Thiruvengadam et al, 2014b). Suitable type of culture medium was helpful for cell/organ growth and also secondary metabolite production (Gai et al, 2015). In the present study, different media, full and half strength MS, B5, NN, and LS were employed in hairy root culture and the results shown that MS medium was superior for biomass accumulation (Table 1).…”

“…The virD2 gene, located outside the T-DNA, is diagnostic for the presence of any remaining Agrobacteria in the root tissue. The integration of Ri T-DNA into the genome of plant cells caused the formation of hairy roots, in which rol and aux genes were harbored (Gai et al, 2015). The rol and aux genes are essential for the induction of hairy roots and it act as a potential activator of secondary metabolites in several plants (Bulgakov, 2008;Park et al, 2011;Nagella et al, 2013;Gai et al, 2015).…”

Remarkable enhancement of flavonoid production in a co-cultivation system of Isatis tinctoria L. hairy root cultures and immobilized Aspergillus niger

“…In this context, we have established a reliable plant in vitro culture system, i.e. I. tinctoria hairy root cultures (ITHRCs), that could supersede the field-harvested Radix isatidis for the efficient production of bioactive flavonoids (Gai et al, 2015).…”

Produção De Raízes Transformadas De Pimenta-Longa Por Infecção Com Agrobacterium Rhizogenes

“…Métodos alternativos para a produção de metabólitos têm sido estudados, e a cultura de tecidos e órgãos vegetais se tornaram ferramentas de grande funcionalidade nesse processo (MORAIS et al, 2012;FILOVÁ, 2014;GAI et al, 2015). Além da cultura de células em suspensão, muitas outras técnicas foram desenvolvidas e estudadas para produção de compostos alvo, como o cultivo de calos e parte aérea, bem como o co-cultivo desses diferentes órgãos e tecidos vegetais.…”