scholarly journals High-efficient transgenic hairy roots induction in chicory: re-dawn of a traditional herb

2016 ◽  
Vol 107 (2) ◽  
pp. 321 ◽  
Author(s):  
Sara Kabirnataj ◽  
Ghorbanali Nematzadeh ◽  
Jafar Zolala ◽  
Ahmad Farhad Talebi
Keyword(s):  
Hairy Root ◽  
Hairy Roots ◽  
Biomass Accumulation ◽  
Ms Medium ◽  
Heterologous Proteins ◽  
Bacterial Strains ◽  
Root Cells ◽  
Whole Process ◽  
Cell Factories ◽  
High Efficient

<p>Plant roots can be manipulated by <em>Agrobacterium rhizogenes</em> to stimulate the production of heterologous proteins for pharmaceutical applications as green cell-factories. During the present study, four bacterial strains (A4, ATCC15834, ATCC11325 and A13) in combination with three co-cultivation media (MS, B5, LS) were examined to establish an efficient and reliable transformation system for chicory (<em>Cichorium intybus</em> L.) using <em>A. rhizogenes</em>. The maximum chicory hairy roots induction was achieved using A13 strain. The observation confirmed that MS medium was more effective on hairy root growth. Dried biomass accumulation of hairy roots infected by A13 strain was 1.10 g l<sup>-1</sup> in MS medium which was significantly higher than those grown in LS and B5 medium (0.88 and 0.72 g l<sup>-1</sup>, respectively). Beta-glucuronidase (GUS) gene was introduced by A13 strain carrying the pCAMBIA1304 binary vector. The results showed that the highest frequency of transformation (63.15 %) was achieved using A13 strain and MS cultivation medium. Detection of GUS and <em>hpt</em>II genes by PCR and GUS histochemical localization confirmed the integrative transformation in hairy roots. In conclusion, the whole process was successfully optimized as a pre-step to manipulate the chicory hairy root cells to improve the unique potential of secondary metabolite production.</p>

scholarly journals First Report of Agrobacterium rubi and A. rhizogenes Causing Crown and Root Gall and Hairy Root on Blueberry in Argentina

Plant Disease ◽  
2010 ◽  
Vol 94 (8) ◽  
pp. 1064-1064 ◽  
Author(s):  
A. M. Alippi ◽  
A. C. Lopez ◽  
P. A. Balatti
Keyword(s):  
Hairy Root ◽  
Hairy Roots ◽  
Crown Gall ◽  
Pathogenic Bacteria ◽  
Vaccinium Corymbosum ◽  
Control Treatment ◽  
23S Rrna ◽  
Bacterial Strains ◽  
First Report ◽  
Multiplex Pcr Assay

From 2006 to 2009, crown gall and hairy root symptoms were observed on blueberry (Vaccinium corymbosum cvs. O'Neil, Millennia, and Misty) plants from six nurseries in Tucumán, Concordia, Pilar, Morón, and Baradero, Argentina. Bacteria were isolated from galls of all three cultivars and from hairy roots of Millenia and O'Neil onto D1 and D1M agar media at 27°C. Typical Agrobacterium colonies developed in 5 days (2). Seven bacterial strains (five from galls and two from hairy roots) were studied further. All were gram negative, aerobic, and catalase positive with rod-shaped cells that synthesized β–galactosidase and metabolized D-glucose, D-arabinose, n-acetyl-glucosamine, maltose, mannitol, and malonate. Strains were negative for lysine decarboxylase, H2S production, indole, and 3-ketolactose production. While gall strains were urease positive and citrate variable (mostly positive), hairy root strains were urease negative, citrate positive, had poly-β-hydroxybutyrate inclusion granules, and clarified acid on potato dextrose agar containing 0.5% CaCO3 (2). Agrobacterium tumefaciens ATCC 15955 and LBA 958 were included as controls. PCR with virA/C primers amplified a 338-bp product corresponding to the virD2 operon and confirmed that the strains harbored a pathogenic plasmid (1). Bacterial strains were assigned to biovars with a multiplex PCR assay targeting 23S rRNA sequences (3). Two strains produced PCR amplicons typical of A. rhizogenes bv. 2. The other five strains produced PCR amplicons typical of A. rubi, which were insensitive to agrocin in a bioassay with A. radiobacter strain K1026. Identity was confirmed by sequencing the 16S rDNA of strains F 266 (GenBank No. GU580894) and F 289 (No. GU580895), which had 99% homology to 16sRNA sequences of A. rubi ICMP 11833 (AY626395.1) and A. rhizogenes ATCC 11325 (AY945955.1), respectively. Pathogenicity of all seven strains was tested on V. corymbosum cv. Misty, Bryophyllum daigremontiana, tobacco cv. Xanthi, tomato cv. Presto, and pepper cv. California Wonder. Plants were inoculated by a needle stabbed into the stems with the appropriate cell suspension (108 CFU/ml) of each strain or with sterile distilled water (control treatment). Two plants of each species were tested per strain. Plants were grown for at least 45 days at 23 ± 3°C and symptoms were recorded. Inoculations with the five strains isolated from galls caused development of spherical, white to flesh-colored, rough, spongy wart-like galls at the inoculation sites. Root strains induced root proliferation on all inoculated plants as well as in a carrot disk bioassay (4). On blueberry plants, galls were dark brown to black, rough, and woody 6 months after inoculation. No lesions were observed on control plants. Bacteria were reisolated from symptomatic tissues of inoculated plants. Enterobacterial repetitive intergeneric consensus-PCR confirmed that the DNA fingerprints of the reisolated strains were identical to those of the original strains. To our knowledge, this is the first report of A. rubi and A. rhizogenes causing hairy root and crown gall on blueberry in Argentina. References: (1) J. H. Haas et. al. Appl. Environ. Microbiol. 61:2879,1995. (2) L. W. Moore et al. Page 17 in: Laboratory Guide for Identification of Plant Pathogenic Bacteria. 3rd ed. N. W. Schaad et al., eds. The American Phytopathological Society, St. Paul, MN, 2001. (3) J. Pulawska et al. Syst. Appl. Microbiol. 29:470, 2006. (4) M. H. Ryder et al. Plant Physiol. 77:215, 1985.

scholarly journals Methyl jasmonate enhances ursolic, oleanolic and rosmarinic acid production and sucrose induced biomass accumulation, in hairy roots of Lepechinia caulescens

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e11279
Author(s):  
Victor M. Vergara-Martínez ◽  
Samuel E. Estrada-Soto ◽  
Susana Valencia-Díaz ◽  
Karlina Garcia-Sosa ◽  
Luis Manuel Peña-Rodríguez ◽  
...  
Keyword(s):  
Methyl Jasmonate ◽  
Bioactive Compounds ◽  
Hairy Root ◽  
Hairy Roots ◽  
Leaf Explants ◽  
Biomass Accumulation ◽  
Wild Plants ◽  
Bioactive Metabolites ◽  
Hairy Root Cultures ◽  
Root Cultures

Background Ursolic (UA), oleanolic (OA) and rosmarinic (RA) acids are bioactive metabolites found in Lepechinia caulescens that have generated interest for their health benefits, which include antimicrobial, antioxidant, antimutagenic, gastroprotective, antidiabetic, antihypertensive and anti-inflammatory properties, among others. To date, very few attempts have been made to evaluate the potential for simultaneous production of these bioactive compounds, using a biotechnological approach. Hairy root cultures offer a biotechnology approach that can be used to study the factors affecting the biosynthesis and the production of UA, OA and RA. In the current study, we established hairy root cultures of L. caulescens and evaluated the effect of sucrose on biomass accumulation, and the effect of different concentrations and times of exposure of methyl jasmonate (MeJA), on the accumulation of UA, OA and RA. Methods Leaves from plants of L. caulescens were inoculated with Agrobacterium rhizogenes strain ATCC 15834. PCR of rolB gene confirmed the transgenic nature of hairy roots. Hairy roots were subcultured in semisolid MSB5 medium, supplemented with 15, 30, 45 or 60 g/L sucrose and after 4 weeks, dry weight was determined. The accumulation of UA, OA and RA of wild plants and hairy roots were determined by HPLC. Finally, the hairy roots were treated with 0, 100, 200 and 300 µM of MeJA and the content of bioactive compounds was analyzed, after 24, 48 and 72 h. Results High frequency transformation (75%) was achieved, using leaf explants from axenic seedlings, infected with A. rhizogenes. The hairy roots showed an enhanced linear biomass accumulation, in response to the increase in sucrose concentration. The hairy root cultures in MSB5 medium, supplemented with 45 g/L sucrose, were capable to synthesizing UA (0.29 ± 0.00 mg/g DW), OA (0.57 ± 0.00 mg/g DW) and RA (41.66 ± 0.31 mg/g DW), about two, seven and three times more, respectively, than in roots from wild plants. Elicitation time and concentration of MeJA resulted in significant enhancement in the production of UA, OA and RA, with treatments elicited for 24 h, with a concentration of 300 µM of MeJA, exhibiting greatest accumulation. Conclusion This is the first report on development of hairy root cultures of L. caulescens. Future studies should aim towards further improving triterpenes and polyphenolic compound production in hairy roots of L. caulescens, for use in the pharmaceutical and biotechnological industry.

scholarly journals Effect of Inoculum Density and Different Media on the Growth of Hairy Roots and Production of Withanolide-A from Withaniasomnifera

2017 ◽  
Vol 16 (4) ◽  
pp. 13-22 ◽  
Author(s):  
Praveen Nagella ◽  
H N Murthy
Keyword(s):  
Medicinal Plants ◽  
Hairy Roots ◽  
Biomass Accumulation ◽  
Inoculum Size ◽  
Inoculum Density ◽  
Ms Medium ◽  
Indian Ginseng ◽  
Crude Drug ◽  
Withanolide A

Withaniasomnifera (L.) Dunal. (Indian ginseng) is one of the most important medicinal plants used as a crude drug for its preventive and therapeutic purposes. Among the diverse constituents of Withania, withanolides are found to be the major components responsible for their biological and pharmacological actions. The present study deals with the effect of inoculum density and different media on the growth of hairy roots and withanolide-A production from Withaniasomnifera. An inoculum size of 10 g/L FW favoured the biomass accumulation(120.42 g/L of FW and 11.98 g/L DW) and withanolide-A production(11.96 mg/g DW) inthe tested range of 2.5, 5.0, 10.0 and 20.0 g/L FW. Among different media tested [Murashige and Skoog (MS), Gamborg’s (B5), Nitsch and Nitsch (NN) and Chu’s (N6)], MS medium favoured both biomass accumulation (121.15 g/L FW and 11.96 g/L DW) and withanolide-A production (11.50 mg/g DW).

scholarly journals Establishment of Hairy Root Cultures ofRhaponticum carthamoides(Willd.) Iljin for the Production of Biomass and Caffeic Acid Derivatives

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Ewa Skała ◽  
Agnieszka Kicel ◽  
Monika A. Olszewska ◽  
Anna K. Kiss ◽  
Halina Wysokińska
Keyword(s):  
Hairy Root ◽  
Hairy Roots ◽  
Leaf Explants ◽  
Biomass Accumulation ◽  
Flavonoid Glycosides ◽  
Transformed Roots ◽  
Caffeoylquinic Acids ◽  
Best Response ◽  
Hairy Root Line ◽  
Half Strength

The aim of the study was to obtain transformed roots ofRhaponticum carthamoidesand evaluate their phytochemical profile. Hairy roots were induced from leaf explants by the transformation ofAgrobacterium rhizogenesstrains A4 and ATCC 15834. The best response (43%) was achieved by infection with A4 strain. The effects of different liquid media (WPM, B5, SH) with full and half-strength concentrations of macro- and micronutrients on biomass accumulation of the best grown hairy root line (RC3) at two different lighting conditions (light or dark) were investigated. The highest biomass (93 g L−1of the fresh weight after 35 days) was obtained in WPM medium under periodic light. UPLC-PDA-ESI-MS3and HPLC-PDA analyses of 80% aqueous methanol extracts from the obtained hairy roots revealed the presence of eleven caffeoylquinic acids and their derivatives and five flavonoid glycosides. The production of caffeoylquinic acids and their derivatives was elevated in hairy roots grown in the light. Only light-grown hairy roots demonstrated the capability for the biosynthesis of such flavonoid glycosides as quercetagetin, quercetin, luteolin, and patuletin hexosides. Chlorogenic acid, 3,5-di-O-caffeoylquinic acid and a tentatively identified tricaffeoylquinic acid derivative were detected as the major compounds present in the transformed roots.

scholarly journals Carrot hairy roots: factories for secondary metabolite production

2020 ◽  
Vol 71 (22) ◽  
pp. 6861-6864
Author(s):  
María A Pedreño ◽  
Lorena Almagro
Keyword(s):  
Phenolic Compounds ◽  
Secondary Metabolite ◽  
Hairy Root ◽  
Hairy Roots ◽  
Hairy Root Cultures ◽  
Root Cultures ◽  
Black Carrot ◽  
Metabolite Production ◽  
Experimental Botany ◽  
Induced Changes

This article comments on: Barba-Espín G, Chen S-T, Agnolet S, Hegelund JN, Stanstrup J, Christensen JH, Müller R, Lütken H. 2020. Ethephon-induced changes in antioxidants and phenolic compounds in anthocyanin-producing black carrot hairy root cultures. Journal of Experimental Botany 71, 7030–7045.

scholarly journals Resveratrol Biosynthesis in Hairy Root Cultures of Tan and Purple Seed Coat Peanuts

Agronomy ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 975
Author(s):  
Ye-Eun Park ◽  
Chang-Ha Park ◽  
Hyeon-Ji Yeo ◽  
Yong-Suk Chung ◽  
Sang-Un Park
Keyword(s):  
Arachis Hypogaea ◽  
Hairy Root ◽  
Hairy Roots ◽  
Seed Coat ◽  
Biological Properties ◽  
Fresh Weight ◽  
Good Source ◽  
Plant Parts ◽  
Seedling Roots

Peanut (Arachis hypogaea) is a crop that can produce resveratrol, a compound with various biological properties, such as those that exert antioxidant, anticancer, and anti-inflammatory effects. In this study, trans-resveratrol was detected in the roots, leaves, and stems of tan and purple seed coat peanuts (Arachis hypogaea) cultivated in a growth chamber. Both cultivars showed higher levels of resveratrol in the roots than the other plant parts. Thus, both cultivars were inoculated with Agrobacterium rhizogenes, in vitro, to promote hairy root development, thereby producing enhanced levels of t-resveratrol. After 1 month of culture, hairy roots from the two cultivars showed higher levels of fresh weight than those of seedling roots. Furthermore, both cultivars contained higher t-resveratrol levels than those of their seedling roots (6.88 ± 0.21 mg/g and 28.07 ± 0.46 mg/g, respectively); however, purple seed coat peanut hairy roots contained higher t-resveratrol levels than those of tan seed coat peanut hairy roots, ranging from 70.16 to 166.76 mg/g and from 46.61 to 54.31 mg/g, respectively. The findings of this study indicate that peanut hairy roots could be a good source for t-resveratrol production due to their rapid growth, high biomass, and substantial amount of resveratrol.

A reliable and efficient protocol for induction of hairy roots in Agastache foeniculum

Biologia ◽  
2014 ◽  
Vol 69 (7) ◽  
Author(s):  
Elnaz Nourozi ◽  
Bahman Hosseini ◽  
Abbas Hassani
Keyword(s):  
Salicylic Acid ◽  
Agrobacterium Rhizogenes ◽  
Hairy Root ◽  
Hairy Roots ◽  
High Performance ◽  
Root Culture ◽  
Hairy Root Culture ◽  
Biochemical Properties ◽  
Separate Experiment ◽  
Transformed Roots

AbstractHairy root culture system is a valuable tool to study the characteristics of gene expression, gene function, root biology, biochemical properties and biosynthesis pathways of secondary metabolites. In the present study, hairy roots were established in Anise hyssop (Agastache foeniculum) via Agrobacterium rhizogenes. Three strains of Agrobacterium rhizogenes (A4, A7 and 9435), were used for induction of hairy roots in four various explants (hypocotyl, cotyledon, one-month-old leaf and five-month-old leaf) of Anise hyssop. The highest frequency of transformation was achieved using A4 strain in one-month-old leaves (51.1%). The transgenic states of hairy root lines were confirmed by PCR (Polymerase chain reaction) method. High performance liquid chromatography analysis revealed that the production of rosmarinic acid (RA) in transformed roots of A. foeniculum was almost 4-fold higher than that of the non-transformed roots. In a separate experiment, hairy roots obtained from one-month-old leaves inoculated with A4 strain, were grown in liquid medium and the effects of different concentrations of salicylic acid (0.0, 0.01, 0.1 and 1 mM) and chitosan (0, 50, 100 and 150 mg L−1) (as elicitor) and sucrose (20, 30, 40 and 50 g L−1) on the growth of hairy roots were evaluated. The results showed that, 30 g L−1 sucrose and 100 mg L−1 chitosan increased the biomass of hairy root cultures and application of salicylic acid reduced the growth of hairy roots compared with control roots.

HAIRY ROOT TRANSFORMATION SYSTEM IN LARGE-SEEDED GRAIN LEGUMES

1995 ◽  
Vol 43 (1) ◽  
pp. 1-5 ◽  
Author(s):  
H.J. Siefkes-Boer ◽  
M.J. Noonan ◽  
D.W. Bullock ◽  
A.J. Conner
Keyword(s):  
Vicia Faba ◽  
Hairy Root ◽  
Hairy Roots ◽  
Faba Bean ◽  
Grain Legumes ◽  
Transformation System ◽  
Root Transformation ◽  
Specific Strain ◽  
Size And Morphology ◽  
Root Size

Hairy roots were produced on faba bean (Vicia faba L.) and chickpea (Cicer arietinum L.) plants by inoculation with Agrobacterium root-inducing strains. Examination of 14 plant genotypes and eight Agrobacterium strains in all possible combinations revealed specific strain/genotype interactions. Hairy root size and morphology differed substantially between faba bean and chickpea hairy roots. Sixty percent of chickpea hairy roots were 10–15 mm in length and forty percent, 15–25 mm. All were <1.0 mm in thickness. Sixty-three percent of faba bean hairy roots were 15–25 mm long and thirty-seven percent, 25–40 mm. All faba bean hairy roots were between 1.0 and 1.5 mm in thickness.

scholarly journals Diazotrophic bacteria in the growth of micropropagated ornamental pineapple

2019 ◽  
Vol 13 (2) ◽  
pp. 269-278
Author(s):  
Adriano Bortolotti Silva ◽  
Ligiane Aparecida Florentino ◽  
Dalvana De Sousa Pereira ◽  
Paulo Roberto Correa Landgraf ◽  
Ana Carolina Rodrigues Alves ◽  
...  
Keyword(s):  
Shoot Growth ◽  
Ex Vivo ◽  
Control Treatment ◽  
In Vitro Cultivation ◽  
Ms Medium ◽  
Ex Vitro ◽  
Diazotrophic Bacteria ◽  
Bacterial Strains ◽  
Indole 3 Acetic Acid

Ornamental pineapple is a hardy plant with significant landscaping value. Tissue culture of plants is viable for producing plants with a high phytosanitary quality. However, one of the difficulties with this cultivar is the acclimatization process, which is slow and can cause losses. The objective of the present study was to verify the potential of inoculation with diazotrophic bacteria for in vitro and ex vivo growth of ornamental pineapple. A group of diazotrophic bacterial strains selected at the Universidade José do Rosário Vellano (UNIFENAS) was prioritized in this study, and the treatments included bacterial strains UNIFENAS (100-13, 100-60, 100-68, 100-153, 100-167 and 100-198). These strains were evaluated in terms of their capacity to produce indole 3-acetic acid. Subsequently, plants were cultivated in a medium composed of MS medium salts (1/4), adding 1 mL of the bacterial strain. In the control treatment, the plants were maintained in 2 mL of MS medium. 7 days after inoculation, the plants were transplanted into the MS, where they were maintained for 30 days. After in vitro cultivation, the plants were transferred to pots containing commercial Plantmax® substrate and maintained under these conditions for 60 days. The diazotrophic bacteria were able to synthesize auxins, and their inoculation promoted greater growth in vitro and ex vitro in the plants. In the acclimatization phase, the plants inoculated with UNIFENAS strains (100-60, 100-68 and 100-153) promoted a higher shoot growth, chlorophyll content and nitrate reductase enzyme activity.

Technology and Application of Hairy Root Culture in Monocotyledons

2021 ◽  
Vol 07 ◽  
Author(s):  
Chang-Qi Hao ◽  
Shuai-Run Wang ◽  
Yi Wang ◽  
Xin-Yi Hou ◽  
Ya-Xuan Jiang ◽  
...  
Keyword(s):  
Medicinal Plants ◽  
Agrobacterium Rhizogenes ◽  
Hairy Root ◽  
Hairy Roots ◽  
Root Culture ◽  
Hairy Root Culture ◽  
Food Crops ◽  
Root Induction ◽  
Hairy Root Induction ◽  
Selection Of

Background: Hairy root culture has been widely used in the production of metabolites in dicotyledons, and a large number of food crops and medicinal plants in monocotyledons need to be developed, but there are many difficulties in the induction of hairy roots in monocotyledons. The purpose of this paper is to introduce the inducing methods, influencing factors and application of hairy roots in monocotyledons, and to promote the development of hairy root system in monocotyledons. Methods: The mechanism of action of Agrobacterium rhizogenes and the current situation of hairy root induction, induction methods and influencing factors of monocotyledons were summarized so as to provide convenience for efficient acquisition of hairy root of monocotyledons. Results: Monocotyledons are not easy to produce phenols, cells are prone to lignification, adverse differentiation and selective response to Agrobacterium rhizogenes strains. It is proposed that before induction, plant varieties and explants should be selected, and different infection strains should be screened. In the process of hairy root induction, exogenous inducers such as acetosyringone can be added. Although these factors can provide some help for the induction of hairy roots in monocotyledons, we still need to pay attention to the disadvantages of monocotyledons from dicotyledons at the cellular level. Conclusion: A large number of food crops and medicinal plants are monocotyledons. Hairy root culture can be used to help the breeding and production of medicinal substances. Therefore, it is necessary to pay attention to the selection of varieties and explants, the selection of Agrobacterium rhizogenes and the addition of acetosyringone in the process of hairy root induction so as to improve the production efficiency and facilitate the development and utilization of monocotyledons.

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