Enhanced production of withaferin A in hairy root cultures of underutilised plant Physalis minima L (original) (raw)
Related papers
2020
Withania somnifera (L.) Dunal is an important ayurvedic medicinal plant used in the traditional Indian pharmacopeia. Withaferin-A is a significant bioactive compound present in the roots of this plant, and it possesses inhibitory effect against the carcinoma cell line in breast cancer, prostate cancer, and liver cancer. The present protocol reveals an efficient hairy root culture system of W. somnifera for withaferin-A production. Leaf explants obtained from in vitro raised plantlets were infected with A. rhizogenes R1000 strain using sonication and heat treatment. Integration of rol B and rol C genes in transformed roots was confirmed by PCR analysis. Elicitation of hairy roots with chitosan has been used for enhancement of withaferin-A production. The accumulation of withaferin-A in hairy root culture has been analyzed using high-performance liquid chromatography (HPLC).
3 Biotech, 2015
An improved Agrobacterium rhizogenes-mediated genetic transformation of Withania somnifera (L.) Dunal was developed using the bacterial strain R1000 with leaf segment explants of in vitro raised plantlets. Out of the three strains used (R1000, MTCC 2364 and MTCC 532), the strain R1000 proved to be more efficient than others. Among the different conditions tested, the highest (93.3 %) transformation rate was observed after 3 weeks when the explants were subjected to sonication (15 s) and heat treatment (41°C for 5 min). Transgenic status of the hairy roots was confirmed by PCR using rol B-specific primers. HPLC analysis showed the ability of hairy roots to synthesize withaferin A and withanolide A, both steroidal lactones of medicinal value. This protocol offers new avenue in A. rhizogenes-mediated hairy root induction and is useful for large-scale production of these bioactive compounds from W. somnifera.
Natural Product Communications, 2018
Withanolides (WTDs), well-known medicinally important compounds of Withania somnifera, including the anticancer compound withaferin A (WFA), are biosynthesized from their dedicated precursor squalene. Condensation of farnesyl pyrophosphate (FPP) molecules to produce squalene is catalyzed by squalene synthase (SQS). In the present study, the Arabidopsis thaliana squalene synthase gene ( AtSQS1) was transformed in W. somnifera by Agrobacterium tumefaciens C58C1 (pRiA4). The AtSQS1 gene was detected in 87.88 % of induced transformed hairy roots (THRs). The results showed that the growth index (GI) of THRs after five weeks of culture was 2-fold higher than that of adventitious hairy roots (AHRs). The biomass doubling time of THRs and AHRs was 18 and 30 days, respectively. Quantitative evaluation of WFA in the studied roots showed that THRs contain 1.51-fold more WFA (330±0.87μg g−1 dry weight (DW)) than AHRs (218±0.17μg g−1 DW). These findings can be used for the production of WFA as a ...
Jamal Academic Research Journal : An Interdisciplinary, 2020
Withania somnifera (L.) Dunal is an important ayurvedic medicinal plant and its primary bioactive compound, withaferin-A possess several pharmaceutical and therapeutic values ranging from immunomodulation to anti-cancer property. For commercial production of withaferin-A, field grown plant materials have generally been used but the quality and uniformity in the levels of active constituents are highly affected by genotype and environmental conditions. Plant tissue culture methods are best alternative way to enhance commercial production of withaferin-A. In this present study, we summarize our recent findings on characterization and screening of wild accessions of W. somnifera collected from Tamil Nadu state. Further, an improved Agrobacterium rhizogenes mediated hairy root culture system was developed and this system providing an efficient tool attaining better transformation efficiency and useful for commercial in vitro production of withanolides.
Plant Cell Tissue and Organ Culture, 2011
A micropropagation system through leaf explant culture has been developed for Withania coagulans. Shoot bud proliferation occurred through both adventitious and de novo routes depending on the hormonal regime of the culture medium. Green compact nodular organogenic callus developed on Murashige and Skoog (MS) medium supplemented with 2.3 μM kinetin (Kn) and lower levels of 6–benzyladenine (BA) (13.3 μM) while multiple adventitious shoot bud differentiation occurred on medium fortified with 2.3 μM kinetin (Kn) and higher levels of BA (22.2 μM). Shoot buds were transferred to proliferation medium containing 2.2 μM BA, 2.3 μM Kn, and 3.9 μM phloroglucinol (PG) for further growth and development of shoot system. Elongated shoots were rooted using a two-step procedure involving pulse treatment of 7 days in a medium containing 71.6 μM choline chloride (CC) and 3.9 μM PG and then transferred to rooting medium containing ½ MS, 1.2 μM IBA, 3.6 μM PAA, and 14.3 μM CC for 3 weeks. Well-rooted plants were transferred to a greenhouse for hardening and further growth. Random amplification of polymorphic DNA (RAPD) showed monomorphic bands in all the plants thereby confirming clonality of the regenerants. Thin layer chromatography (TLC) showed the presence of withanolides in the regenerated plants. Quantification through reverse-phase HPLC revealed increased concentration of withanolides in the regenerated plants compared to the field-grown mother plant. Accumulation of withaferin A and withanolide A increased up to twofold and that of withanone up to tenfold. Direct regeneration via leaf explants will be useful for Agrobacterium-mediated genetic transformation, and will facilitate pathway manipulation using metabolic engineering for bioactive withanolides.
Journal of Applied Biotechnology & Bioengineering
Withania somnifera (L.) Dunal is an important medicinal plant with many pharmaceutical and therapeutic uses ranging from immunomodulation to anticancer property. Field cultivation of W. somnifera is time consuming and it not able to meet current global demands. For commercial production of withanolides, field grown plant materials have generally been used but the quality and uniformity in the levels of active constituents are highly affected by genotype and environmental conditions. In vitro culture methods are the best alternative and have possible ways to enhance commercial production of withanolides. In this review, we summarize our recent findings on characterization and screening of wild accessions of W. somnifera collected from Tamil Nadu state. Further, an improved Agrobacterium rhizogenes mediated hairy root culture system was developed and this system providing an efficient tool attaining better transformation efficiency and useful for commercial in vitro production of withanolides.
Withania somnifera (Ashwagandha) is a plant used in medicine from the time of presence by ayurveda, the ancient system of Indian medicine and in plant biotechnology. The dried roots and leaves of the plant are used in the treatment of various nervous and sexual disorders. Medicinal properties of plant have been attributed to the presence of a group of steroidal lactones which are known as withanolides which are present in roots and leaves. Withaferin A is a steroidal lactone being derived from Withania somnifera. It is used in ayurvedic medicine. It has a wide range of activities which comes under pharmacy ,cardio protection, the anti-inflammatory agent , an immuno modulatory, an anti-angiogenesis, anti-metastasis and anti-carcinogenic property. This review article is focusing on the methodology to achieve the maximum production of withaferin A employing cell suspension culture of Withania somnifera. As withaferin A low concentration was found to be a limiting factor in previous reports on Withania somnifera. This article will encompass the parameters which are responsible for optimum production of withaferin A employing cell suspension culture of Withania somnifera.
Deleted Journal, 2020
Plants are a tremendous source for the discovery of new products with medicinal importance in drug development. Several distinct chemicals derived from plants are used in various important ways. Secondary metabolites are economically important as drugs, flavor, dye, pesticides, and food additives. Plants produce the diversity of secondary metabolites which not only plays an important role in adaption according to the environment but also represents an important source of active pharmaceuticals. The possibility of altering the production of bioactive plant metabolites through tissue culture technology is one of the emerging fields of biotechnology to investigate and enhance the production of secondary metabolites. This enhancement through field cultivation has many defects such as slow growth and low and variable yield due to the environmental and biotic factors. Therefore, hairy root culture has been developed as a more efficient alternative biotechnological tool for secondary metabolite synthesis, regardless of environmental, seasonal, and climatic variations. In vitro hairy roots formed by genetic transformation have been efficiently utilized for the synthesis of higher levels of flavonoids due to their biochemical and genetic stability as well as their fast growth in media without phytohormones. The focus of the present review is a detailed assessment of research on rhizogenesis in different plants using Agrobacterium rhizogenes for the last twelve years particularly for the enhancement of secondary metabolites. The study reveals different techniques involved for rhizogenesis in different plants, compatibility trends of the desired gene, and modifications in the techniques during these years.
2014
Genetic transformation was carried out in ashwagandha (Withania somnifera (L.) Dunal) using three different Agrobacterium rhizogenes strains viz., A4, ATCC 15834 and MTCC 2364, for inducing hairy roots. The explants such as hypocotyls, cotyledonary segments, leaf segments, shoot tips and nodal segments were used for genetic transformation. A4 and ATCC 15834 strains produced successful transformation and hairy (transformed) roots were induced from leaf segments and shoot tips. A4 strain produced transformation by direct inoculation of bacteria from single cell colonies as well as in the suspension form, but ATCC 15834 produced transformation only in the suspension form. Among the liquid media tested, half MS was found to be superior in promoting hairy root growth. The transformation was confirmed by PCR and dot blot analysis. A Thin Layer Chromatographic method was employed for withanolide estimation. The spot corresponding to withaferin A was observed under UV at 254 nm. Field root ...