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Role of Tissue Culture Technique on Wheatgrass (Triticum aestivum): A Staple Food Commonly known as Wheatgrass Beneficial for Human Health |
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| Paper Id :
18776 Submission Date :
2024-03-13 Acceptance Date :
2024-03-20 Publication Date :
2024-03-25
This is an open-access research paper/article distributed under the terms of the Creative Commons Attribution 4.0 International, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. DOI:10.5281/zenodo.11076926 For verification of this paper, please visit on
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| Abstract |
Wheatgrass is a type of worldwide acceptable staple food. The scientific name is Triticum aestivum belongs to Poaceae family. The plant is proved to be superfood which has all nutrients and antioxidants. Many Scientists and researchers reveals many positive experiment through tissue culture techniques on Wheatgrass. They are responsible to boost up immune system, aid in digestion, and provide energy. The color of plant is green in colour having healthy shoots and roots. It is regarded as a super potent health food with amazing benefits. In present work, the different types of plant growth regulators were taken. It includes Auxin, Gibberlin, Cytokinin and IBA. These hormones were taken in different concentration like 10-3M, 10-6M and 10-9M, MS media was taken which was supplemented with different concentration of hormones. Here two types of media were used i.e liquid and solid media. The liquid media was supplemented with hormones and MS media, while in solid media both MS and 0.8% Nutrient Agar were used. In liquid media, seed culture was done in Ms media supplemented with hormones. Out of four types of Plant Growth Regulators, Auxin plays a good role in morphogenesis and growth of shoots and roots in both liquid and solid culture. The seed germination started within 48 hours of inoculation. Within one week of short interval of period, the shoots measures 7cm and the length of roots ranges upto 4cm in length. In solid supplemented media, the shoots and roots ranges upto 4cm to 5.2 cm in length in 10-6M Auxin supplemented media. Later, both protocols cultured in solid and liquid media were transfer to sterilized pot provided with soil. The parameters of soil were checked before inoculation of plants in pots. In solid culture, the nutrient Agar arises good results in morphogenesis and growth of roots and shoots of plantlets. The concentration of 10-3 M and 10-9M of Gibberlin, Cytokinin and IAA supplemented media shown a very negligible result on growth and morphogenesis of plantlets. On the contrary to other concentration 10-6 M plus MS media considered a good candidate for developing healthy plantlets. The nature of cultivated in vitro plants of wheatgrass resembles same as the natural plants in morphology but nutrient contents seems to be altered. The objective of this study was to evaluate the role of tissue culture technique on the fruitful production of protocols with the application of Plant Growth Regulators. The nutritional analysis was also done from the plants of Wheatgrass harvested from in vivo and in vitro plants. |
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| Keywords | Wheatgrass, Triticum aestivum, Plant Growth Regulators (Auxin, Gibberlin, Cytokinin and IBA), MS media, Nutrient Agar, Shoots and Roots. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Introduction | Wheatgrass is a type of prominent crops belongs to poaceae family. Many seasonable cereals such as wheat, barley, oat, rye and triticale are acceptable crops for in vitro regeneration. Since a decades, many scientists and researchers involves in regeneration of allied species of crops for producing transgenic plants (Gales et al. 1998). In cereals, the crops are transformed to mature plant with the application of many Biotechnology applications. The use of cell and tissue culture systems has a significant impact to produce disease resistant variety of crops. Mostly cereal regeneration includes induction of embryogenesis calluses from immature tissues and further subsequent regeneration of plantlets (Jimenez et al. 2001). In most of the cases, the process of regeneration is seems to be time-consuming and laborious process. Many literature surveys on somaclonal variation in different types of crops show multitasking results of callus induction and regeneration process (Bennid et al. 1996). In some cases, the abnormal morphological characteristics and reduced fertility were also reported in many cultured crops during Plant tissue Culture Technique using Plant Growth Regulators (Trewavas 1981). In contrast to immature tissue, explants are seems to be most active derived from immature embryos or scutella in wheat in culture medium[1]. In some reports, the required culture and induction of direct shoot regeneration depends on the nature of the plant organ (Ganeshan et al. 2006) which the explant was regenerated. It is highly responsive on character of plant genetic modification (Repellin et al. 2001). The direct shoot proliferation protocol depend on the nature of medium and the type of the explants for elite cultivars of wheat. The activation of shooting response depends on the concentration of cytokinin supplemented in the medium. Cytokinins works as signaling molecules that activate totipotent cells of callus for shoot organogenesis. These molecules activate as perfect machinery of somatic (Eudes et al. 2003) cells (leaf, stem, cotyledon and so on) for direct organogenesis, while of shoot apex they stimulate the growth caused by presence of meristemic cells at the tip of explants. These cytokinin may also shows a positive impact on the explants to produce multiple shooting responses. Taxonomy Kingdom Plantae Division Magnoliophyta Class Liliopsida Order Cyperales Family Poaceae Genus Triticum Species Aestivum Common Wheatgrass Name History
of Wheatgrass (Triticum aestivum)
Wheatgrass has been used in Indian tradition since a decades. The consumption of wheatgrass was first began in western world in the 1930. Later, it spread throughout the western countries like United States and Canada. Dr. Charles F. Schnalad first highlighted the role of the use of Wheatgrass in our diet in 1932. Besides, he also developed a product called “Carophyl” which is known as “World First Multivitamin” (Figure 1d). The glass of wheatgrass in our daily diet also provides beneficial aspects for human health. Besides, multivitamin tablets also have a good impact which provides all vitamins for perfect growth and development of body (Figure 1f & 1g). Since decades, Wheatgrass is traditionally used in both Persian and Indian festivals. Many rituals of Hindu culture are done with cultivating wheatgrass. During Navaratri pooja, Hindu sow wheatgrass seeds on the first day of Navaratri pooja and offer the saplings to the mother goddess on the last day as part of the rituals. |
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| Objective of study |
The aim of this study was to assess the significant role of tissue culture technique with the application of plant growth regulators and nutritional analysis was also done from the plants of Wheatgrass harvested from in vivo and in vitro plants |
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| Review of Literature | The role of micro-propagation has a good link with regeneration and callus morphogenesis (George et al. 2005). The abiotic factors like temperature, salinity, heat, pressure plays a great role on growth and development for expalnts in even aseptic condition. Light condition during in vitro phase had a positive influence on the responses of explants to direct shoot regeneration (Alizadeh et al. 2004). The explants that initially were kept in darkness and then exposure to photoperiod showed better response than those maintained in photoperiod treatment. That is, darkness was observed to stimulate more direct shoots than light condition. These findings were supported by the studies in and on wheat. The effect of light can be a good source of metabolism and sugar uptake. Many scientist and researcher reported a good response of hormones ( Davis et al. 1995) in developing protocol for shoot embryonic meristem with the scutellum, the combination of 10 mgl-1 (BAP) and 2 mgl-1 (2, 4-D) (Fujimura et al. 1980) and darkness for 4 weeks can enable to enhance the rate of plant regeneration (Sharma et al. 2004) and do reduce the number of days required to direct shoot regeneration using in vitro culture. |
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| Methodology |
Literature Survey and Collection of seeds A regular literature survey was done before collection of seeds. A good variety of seeds was brought from nearby seed bank nearby Madhav University Campus, Aburoad Rajasthan. Preparation of MS Media Composition of Murashige and Skoog's (MS) Media (Murashige and Skoog 1962)
Germination of Seeds Here the seeds was first sterilized in CuSo4 Solution, Later they were allowed to germinate on filter paper moisten with nutrient solution i.e MS media having macronutrient, micronutrient and Vitamin solution for 12 hour. After 12 hour, the seeds spread over the filter paper in four individual petriplates. The petriplates were kept at different parameters like light and dark condition. The temperature was maintained at 22 to 250C. These petridish were kept in a thermostatically controlled culture room at 25 + 20C under 12 : 12 light : dark cycle. Media used The germinated seeds inoculated in plant growth regulators (Auxin, Gibberlin, Cytokinin and IBA) supplement Agar plus MS Media, The amount of Plant Growth Regulators was 10-3M, 10-6M and 10-9M. In vitro culture of different types of crops Transfer of In vitro cultured Plants to In vivo condition The process of germination started within 48 hours. When the size of the seedlings reached about 5 cm, the plant pot was transferred to the greenhouse. The temperature of the greenhouse was constantly maintained at 25 ± 20C, and 900 lux of artificial cool fluorescent light with a 14 hour photoperiod was provided to compensate the short day length. The height of mature plants ranged from 115cm after 7 days of inoculation inside the soil. Nutritional and Mineral content was Analysis by AOAC 1990 and Pavel 2009. |
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| Result and Discussion |
Wheatgrass is a major source of nutrients essential for health. It can be cultivated in throughout the year. Wheatgrass can be grown indoors or outdoors. The role of Plant Tissue Culture techniques have a great significant role to fertile the wide verities of crops in short interval of time. In vivo condition, it is not possible to cultivate in all season but in vitro condition, it can grow throughout the year in aseptic condition as we desire.
Figure 1a: A bunch of elongated shoots of Triticum aestivum (Wheatgrass) cultured in MS Nutrient supplement Plant Growth regulator media. Figure 1b: Regenerated Shoots and Roots of Triticum aestivum (Wheatgrass) in MS plus Plant Growth regulators supplemented solid media. Figure 1c: Regenerated shoots transfer to pot for further growth of Triticum aestivum (Wheatgrass) plants. Figure 1d: Juice extractor to prepare the juice of Shoots of Triticum aestivum (Wheatgrass) plants. Figure 1e: A fresh glass of Wheatgrass Juice. Figure 1f: A bottle of Wheatgrass tablets. Figure 1g: A health tonic of Wheatgrass called Carophyl. In in vitro condition, it grows in MS medium with supplementation of Plant Growth Regulators. Leaves are harvested when they develop a "split" as another leaf emerges. George et al (2008) reported a brilliant view on micro-propagation during Tissue culture. In most of the literature, the callus phase is exclusive concession that used in Rice (Nhuts et al. 2000), cereal (Eudes et al. 2003) to decrease the time, spend, frequency of somaclonal variation and especially genotype dependency that is consistent to (Sharma et al. 2004). Similar genotype were developed in present study. Here different types of plant growth regulators were taken. It includes Auxin, Gibberlin, Cytokinin and IBA. These hormones were taken in different concentration like 10-3M, 10-6M and 10-9M, MS media was taken which was supplemented with different concentration of hormones. Here two types of media were used i.e liquid and solid media. The liquid media was supplemented with hormones and MS media, while in solid media both MS and 0.8% Nutrient Agar were used. In liquid media, seed culture was done in Ms media supplemented with hormones. Out of four types of Plant Growth Regulators, Auxin plays a good role in morphogenesis and growth of shoots and roots in both liquid and solid culture. The activation of shooting response depends on the concentration of cytokinin supplemented in the solid medium (Figure 1b). Cytokinin also shows a positive attitude of signaling molecules that activate totipotent cells of callus for shoot organogenesis. These molecules activate somatic cells (Shoots, roots and cotyledon) for direct organogenesis ((Malik et al. 2007, Nhut et al. 2002). The shoots were emerges out and stimulate the growth occur due to meristemic cells at the tip of explants. These cytokinines may also turn the explants to produce multiple shooting responses (Centeno et al. 1996) . At the tip of the plantlets water droplets are observe which shows the sign of Guttation (Figure 1c). Our results reported on the seed germination started within 48 hours of inoculation in 10-6M Auxin supplemented MS media (Figure 1a and 1c). However, within one week of short interval of period, the shoots grow out measures 17cm in and the length of roots ranges upto 5cm in length. The color of shoots was dark green. Similarly, The roots were developed elongated with sharp root hairs, dark brown in color. On the other hand, In solid nutrient Agar supplemented plant growth regulator combination, the shoots and roots ranges The meristem cells has a great achievements in regeneration of plantlets in direct regeneration (Ahmad et al. 2001). The role of micro-propagation has a good link with regeneration and callus morphogenesis (George et al. 2005). The abiotic factors like temperature, salinity, heat, pressure plays a great role on growth and development for expalnts in even aseptic condition. Light condition during in vitro phase had a positive influence on the responses of explants to direct shoot regeneration (Alizadeh et al. 2004). The explants that initially were kept in darkness and then exposure to photoperiod showed better response than those maintained in photoperiod treatment. That is, darkness was observed to stimulate more direct shoots than light condition. These findings were supported by the studies in and on wheat. The effect of light can be a good source of metabolism and sugar uptake. Many scientist and researcher reported a good response of hormones ( Davis et al. 1995) in developing protocol for shoot embryonic meristem with the scutellum, the combination of 10 mgl-1 (BAP) and 2 mgl-1 (2, 4-D) (Fujimura et al. 1980) and darkness for 4 weeks can enable to enhance the rate of plant regeneration (Sharma et al. 2004) and do reduce the number of days required to direct shoot regeneration using in vitro culture. Similar, response was observed in present study with the combination of Auxin, Gibberlin, Cytokinin and IAA plus MS Agar medium, lastly it has been reported that none of the regenerated plantlets in this study showed morphological abnormalities and set seeds normally. The leaves were erect except for the oldest two or three leaves. The tillers of the five 17 plants continuously supplied panicles at different developmental stages for this experiment. The panicles not used for the experiment developed to maturity. The number of florets on a panicle was estimated to be about 60 to 110. From randomly selected panicles, the rate of sterility was determined to be 84 percent for the earlier panicles. Seed sterility was significantly decreased as subsequent tillers matured to produce their panicles. The growing season and conditions and/or lack of wind to aid self-pollination in the greenhouse was most likely responsible for the high sterility of the plants. The seeds observed from this experiment was good and healthy. Nutritional and Mineral content Triticum aestivum is commonly known as Wheatgrass. It is a major stable food in everyone diet. It has rich nutritional components. In comparison to other land plants, wheatgrass also contain a high amount of chlorophyll a, b , c , antioxidant and enzymes. Many researcher and scientist reported the health benefits of wheatgrass in human life. It provides a major source of supplemented nutrition. It is a source of minerals like potassium, calcium, copper, iron, zinc, copper, manganese, selenium etc. Besides, Wheatgrass also contain protein, dietary fiber, vitamin A, vitamin C, vitamin E, vitamin K, Vitamin B, Vitamin B2, Vitamin 6 etc. In contrast, It is also a good source of protein . Khan et al. (1984) has discuss the role of nutritional components of wheatgrass. In his report, positive attribute of nutritional components was engulfed and prescribed to add the wheatgrass products in everyday life. (Table 1). Figure 2 shown a very sharp amount of mineral and other nutritional contents present in wheatgrass. The composition of nutritional component seems to fluctuate. The juice of wheatgrass provides all nutrients necessary for body building biological activity. The nutrient content of wheatgrass juice is almost roughly equivalent to that of dark leafy vegetables. Pavel et al. 2009 reported the techniques on analysis of chemical and nutritional components of different types of crops. Many studies proved the wheatgrass to have freshly sprouted first leaves. They are used as a food, health drink, or dietary supplement. The juice of wheatgrass juice has a best complements to have a best ingredients in all juice bars to fulfill the body fitness. Due to its high content of bioactive compounds, it could be recommended for consumption as a fresh juice blend to elevate fertility and promote youthfulness. This juice meets the changes in the present-day consumers’ diet chart. It becomes a vital change in the marketing trends of the food sector. It also highly recommended the consumption of juices because of the importance for micronutrient and macronutrient (Figure 1d & 1e). Wheatgrass is available easily in most part of the countries as fresh as produce, in tablets, frozen juice, shakes with fruits, mints, coriander and powder. Besides above, Wheatgrass is also famous in commercial market as a cosmetic, foliar spray, face packs, creams, hair gel, massage lotion, fertilizer and liquid herbal supplement.
Figure2 : Graphical representation of Nutritional and Mineral components of Wheatgrass. |
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| Conclusion |
The conclusion of this study was to evaluate the role of tissue culture technique on the fruitful production of protocols with the application of Plant Growth Regulators. The use of culture media shows a positive response in the production of thousand protocols within a short interval of time. Besides, The nutritional analysis was also done from the plants of Wheatgrass harvested from in vivo and in vitro plants. In vitro shoots shows a better impact of nutrients in comparison to in vivo plants. Thus wheatgrass plays a prominent role for health and scientific temperament. |
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| References | 1. Ahmad A., H. Zhong, W.L. Wang and M.B. Sticklen (2001). “Shoot apical meristem: In vitro regeneration and morphogenesis in wheat (Triticum aestivum L.)”, In Vitro Cell Dev. Biol.-Plant, vol 38, pp. 163-167. 2. Alizadeh H., M.R. Naghavi, M. Omidi and B. Saatian (/2004). “Effect of plant growth regulators on direct shoot regeneration of wheat (Triticum aestivum) ”, Australian Agronomy Conference, 12th AAC, 4th ICSC, 2004. 3. AOAC (1990). Official Methods of Analysis of Associationof Analytical Chemist. 15th Edition, WashingtonD.C.: AOAC. 4. Bennici A., P. G. Cionini and F. D'Amato (1976). “ Callus formation from the suspensor of Phaseolus coccineus in hormone-free medium: A cytological and DNA cytophotometric study”, Protoplasma, vol 89, pp. 251-261. 5. Centeno M. L., A. Rodriguez, I. Feito and B. Fernandez (1996). “Relationship between endogenous auxin and cytokinin levels and morphogenic responses in Actinidia deliciosa tissue cultures”, Plant Cell Reports, vol 16, pp. 58-62. 6. Davies P. J.(1995 ).“The plant hormone concept: concentration, sensitivity and transport”, in , Plant Hormones Physiology, Biochemistry and Molecular Biology, 2nd ed., P. J. Davies., Ed. London, UK: Kluwer Academic publishers,pp. . 13–38. 7. Eudes F., S. Acharya, A. Laroche, L. B. Selinger and K. J. Cheng (2003). “A novel method to induce direct somatic embryogenesis, secondary embryogenesis and regeneration of fertile green cereal plants”, Plant Cell, Tissue and Organ Culture, vol 73, pp. 147–157. 8. Fujimura T. and A. Komamine (1980).“Mode of Action of 2, 4-D and Zeatin on Somatic Embryogenesis in a Carrot Cell Suspension Culture”, Zeitschrift Für Pflanzenphysiologie, vol 99, pp. 1-8. 9. George E.F. and P.C. Debergh,( 2008). “Micropropagation: Uses and Methods,” in Plant Propagation by Tissue Culture, 3rd ed., E. F. George, M. A. Hall and G. J. De Klerk., Ed. Dordrecht, The Netherlands: Springer, pp. 29-64. 10. Galess C., H. Lorz and A. Jahne-Gartner. (1998). “Establishment of a highly efficient regeneration system from leaf base segment of oat (Avena sativa L.) ”, Plant Cell Rep, vol 17, pp. 441 -445. 11. Ganeshan S., S.V. Chodaparambil, M. Baga, D.B. Fowler, P. Hucl, B.G. Rossnagel, R.N. Chibbar (2006). “In vitro regeneration of cereals based on multiple shoot induction from mature embryos in response to thidiazuron”, Plant Cell Tissue Organ Cult, vol 85, pp. 63-73. 12. Jimenez V. M. and F. Bangerth (2001). “In Vitro Culture And Endogenous Hormone Levels in Immature Zygotic Embryos, Endosperm and Callus Cultures of Normal and High-Lysine Barley Genotypes”, Journal of Applied Botany, vol 75, pp. 1-7. [11] Khan M. (1984). Nutritional Attributes of Wheat. Birmingham, AL: Progressive Farming. 13. Malik S., M. Zia, R. Rehman and M. Fayyaz Chaudhary (2007). “ In vitro plant regeneration from Direct and indirect organogenesis of Memordica charantia ”, Pakistan Journal of Biological Sciences, vol 10 (22), pp. 4118 -4122. 14. Nhut D. T., B. V. Le and K. Van (2000). “ Somatic embryogenesis and direct shoot regeneration of rice (Oryza sativa L.) using thin cell culture of apical meristemic tissue”, J. Plant Physiol, vol 157, pp. 559-565. [13] Pavel, K. (2009). Chemical composition and nutritional value of European species of wild growing mushrooms: a review. Food Chem.2009, 113(1):9–16 15. Repellin A., M. Baga, P. P. Jauhar and R. N. Chibbar (2001). “Genetic enrichment of cereal crops via alien gene transfer: New challenges”. Plant Cell Tissue Organ Cult, vol 64, pp. 159–183 16. Sharma K., R. Hansch, R. Mendel, and J. Schulz. (2004).“A Highly Efficient Plant regeneration System through Multiple Shoot Differentiation from Commercial Cultivars of Barley (Hordeum Vulgare L.) Using Meristematic Shoot Segments Excised Rom Germinated Mature Embryos”. Plant Cell Rep, vol 23, pp. 9–16. 17. Trewavas A (1981). “ How Do Plant Growth Substances Work? ”, Plant Cell & Environment, vol 4, pp. 203-228. |
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