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Callus formation and shoot regeneration in Aloe vera ; callus induction was developed in MS media with Kn (0.5 mg/L) and 2,4-D (0.5mg/L) and shoot formation with 1mg/L BAP and 0.5 mg/L NAA.
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In vitro regeneration of medicinal plants is important and potential for the production of high quality plant based medicine. Aloe vera belongs to the Liliaceae family and frequently used for medicinal purpose, but its growth rate is very slow naturally because of male sterility. In vitro culture is used for commercial production and is achieved in...
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... mercuric chloride and surface sterilization was done to remove the trace of surface borne microorganism. The shoot tips were cut with the help of scalpels and forceps into 1-2 cm and then again rinsed 2-3 times with distilled water. Media for tissue culture contains all major and minor elements, vitamins and growth regulators which are essential for normal plant growth. MS basal media [12] used for culture of explants contains sucrose (3%) as carbon source, agar-agar (7%) as solidifying agent and PVP (1g/l) which reduces the polyphenolic substances that are supplemented with different concentration of plant growth hormones such as IAA, Kn, BAP (pH of media 5.6-5.8) and finally autoclaved. Explants were inoculated vertically in culture bottles containing 50-70 ml MS medium with 0.2mg/L BA and 0.2 mg/L IBA media, and then culture bottles were kept in growth chamber under 16/8 h (light/dark) condition, temperature was 25±2oC and light intensity 2500-3000 lux; generally white fluorescent lamps were used because blue and red light induces shoot and root formation in many species [13]. Explants were subcultured after 20-25 days on different concentration of plant growth hormones for promoting shoot and root formation. In vitro regeneration of plants was regulated by different concentration of plant growth regulators. After 15-20 days of inoculation explants were swelled and induction of callus started. Since it is a coherent and amorphous tissue, callus were transferred to new MS medium supplemented with BAP 1mg/L and NAA 0.5 mg/L for promoting shoot formation of callus. Generally cytokinins induced shoot bud formation (cytokinin/auxin>1), auxin induced root formation (auxin/cytokinin>1) and when concentration is equal, they tend to induce callus formation. We used different concentration of growth regulators but the best result of callus induction was developed in MS media with Kn: 0.5 mg/L and 2,4-D: 0.5mg/L and shoot formation ( Figure 1) with 1mg/L BAP and 0.5 mg/L NAA.By using different concentrations of BAP (mg/l) with NAA (mg/l) we got different number of shoots at different time point the statistical analysis was done (Table 1). Callus cultures of medicinal plants have been done by many scientists [14,15] with bud initiation due to effect of BA, IAA and NAA [16,17]. When increasing the concentration of BAP in nutrient media (BAP: 1.0 and NAA: 0.5), number of shoots increased per culture. After 3-4 weeks of inoculation, the newly formed shoots were excised in aseptic condition and transferred to new MS media which was supplemented with different concentration of growth hormones. It has been reported that acetic acid also helped in shoot proliferation of Aloe vera [18]. Many plants that are rich in polyphenolic compounds inhibit the primary cultures; such compounds turned the media brown when the explants were cultured [19] due to the oxidation of polyphenolic compounds of explants. A. vera by having such problems, the explants were transferred on fresh medium every two weeks [20,21] and polyvinylpyrrolidone (PVP) was also added to the culture media for successful in vitro ...
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Aloe vera is a succulent belongs to the Liliaceae family. It is a native, chiefly to the warm dry areas of South Africa, but cannot survive freezing temperatures. The cultivation of Aloe vera has acquired great commercial importance for medicinal products and cosmetics processing but information is scarce about processing of this crop. This investi...
Citations
... 1(b) and 2). Combination of BAP and NAA at 1.0 mg/L BAP and 0.50 mg/L NAA, respectively, has resulted in significantly higher mean shoot numbers in many aloes species [7,13,15,16]. However, other studies have reported higher mean shoot number per explant at higher concentrations of BAP [8,12,17,18]. ...
Objective
Aloe elegans Tod. is an ecologically, environmentally, medicinally, and commercially useful aloe species in Ethiopia and Eritrea. Unfortunately, it is highly threatened due to industrial and urban expansion and traditional mining and agricultural activities. As a consequence, it is included in the IUCN List of Threatened Species since 2013. The plant is getting thinly populated in many parts of the Tigrai floristic region since it is being exploited for traditional and commercial purposes. Therefore, this study was aimed to develop a reproducible, large-scale micropropagation protocol using offshoot cuttings in Murashige and Skoog (MS) media enriched with plant growth regulators (PGRs).
Results
Sterilized explants cultured in full-strength MS media enriched with 0.25 mg/L benzyl amino purine (BAP) and 0.10 mg/L naphthaleneacetic acid (NAA) resulted in 100% healthy and live (i.e., initiated) explants after four weeks of initiation study. Unsupplemented initiation media (control) yielded only 14.3% initiated explants. The initiated explants were tested for their shooting response to produce microshoots by incubating in different concentrations and combinations of BAP and NAA for four weeks. Fewer days to shooting (13.0 ± 1.0 days), higher mean shoot number (5.0 ± 1.0), and higher mean shoot length (9.20 ± 2.35 cm) were observed with 1.0/0.50, 1.0/0.25, and 1.0 /0.50 mg/L BAP/NAA combinations, respectively. The rooting responses of the microshoots toward producing plantlets were also tested by incubating them in half-strength MS media enriched with different concentrations of NAA and indole-3-butyric acid (IBA) for four weeks. Fewer mean days to rooting (12.0 ± 1.0 days), higher mean root number (8.0 ± 4.0), and higher mean root length (7.53 ± 3.03 cm) were observed in MS media enriched with 0.75, 0.75, and 1.25 mg/L IBA, respectively. The responses of A. elegans plantlets to primary (in greenhouse) and secondary (in nursery shade and direct sunlight) acclimatization in coco peat, composted soil, and manured soil media were high – with survival percentages of 87.5–97.8% in three to four weeks.
... The discovery and documentation of the role of plant growth regulators (PGRs) like auxins and cytokinins in tissue culture served as major thrust for the crop improvement. Auxins, cytokinins, and auxin-cytokinin interactions are usually considered to be the most important for regulating growth of plant under control condition, as these two classes of hormones generally required for root and shoot regeneration (Choudhary et al., 2011;Ahmad et al., 2020). Root induction and callus proliferation is a dominating characteristic for most of the auxins (Adelberg and Naylor-adelberg 2012). ...
The discovery and documentation of the role of plant growth regulators in tissue culture served as major thrust for the crop improvement. For getting maximum response, utilization of suitable growth regulator at appropriate concentration needs to be carefully balanced and controlled. Therefore, the present investigation was carried out to optimize the levels of Indole-3-acetic acid (IAA) for micropropagation of Gwarpatha [Aloe vera (L.) Burm. f.]. Lateral shoot explant was used for inoculation with incorporation of different concentrations of IAA (0.5-4.0 mg/L) in Murashige and Skoog (MS) medium. Diverse regeneration responses were observed for shoot multiplication, root induction and for callus induction at varied levels of IAA. Highest number of shoot per explant (2.90) was recorded in the treatment of 3.0 mg/L with 70 % induction frequency while, maximum number of root (4.10) was induced at 2.0 mg/L. Moreover, highest callus weight (1.24 g) was reported at 4.0 mg/L with yellow brown colour and semi-compact texture. These identified levels of IAA, further can be utilized for efficient micropropagation of Aloe vera in view of reducing economic cost.
... The discovery and documentation of the role of plant hormones like auxins and cytokinins in plant tissue culture served as major thrust for the crop improvement. Auxins, cytokinins, and auxin-cytokinin interactions are usually considered to be the most important for regulating growth of plant under in vitro condition, as these two classes of hormones generally required for root and shoot regeneration (Choudhary et al., 2011). The NAA (1-naphthalene acetic acid) is one of the important auxin, commonly utilize in plant tissue culture for micropropagation of various species. ...
The NAA (1-naphthalene acetic acid) is an important auxin, commonly utilize in plant tissue culture for micropropagation of various plant species. The degree and direction of regeneration response of explant, highly influences with the concentration of growth regulator. Therefore, in the present investigation, effects of different levels of NAA (0.5 to 4.0 mg/l) were evaluated for shoot proliferation, callus induction and root induction of Aloe vera. Various concentrations of NAA had shown differential effects on lateral shoot explant in MS (Murashige and Skoog) medium. The treatment of 2.5 mg/l induced highest number of shoots (3.10) per explant, followed by 3.0 mg/l NAA (2.90) with 80 per cent induction frequency. Maximum callus weight (1.93 g) was observed at 3.5 mg/l with 50 per cent frequency while, highest number of roots (5.0) was obtained at 1.5 mg/l with root induction frequency of 90 per cent. Thus, for best economical use, these identified levels of NAA can be utilized for getting maximum regeneration response of Aloe under in vitro condition.
... Callus induction and organogenesis in the form of shoot/root/embryogenesis from somatic cells and tissues may occur depending upon the concentration of plant growth regulators such as auxins and cytokinins singly or in combination. Auxins, cytokinins, and auxin-cytokinin interactions are usually considered to be the most important for regulating growth of plant under in vitro condition, as these two classes of hormones generally required for root and shoot regeneration (Choudhary et al., 2011). Two major properties of cytokinins that are useful in culture are stimulation of cell division and release of lateral bud dormancy. ...
The BAP (6-benzylaminopurine) and Kn (Kinetin) are important cytokinins, commonly utilizes in plant tissue culture for micropropagation of various plant species. However, their regeneration response under in vitro condition highly dependent on the culture condition especially concentration of growth regulators. In the present investigation, effects of different levels of BAP and Kn (0.5 to 6.0 mg/l) were evaluated for callus induction in lateral shoot explant of Aloe vera. Differential concentrations of these growth regulators had shown variable effects on callus proliferation in MS (Murashige and Skoog) medium. In BAP incorporated medium, maximum callus induction (0.92 g) was obtained at the concentration of 5.0 mg/l BAP, which was characterized with yellow colour and semi compact texture. However, highest callus weight induced in Kn fortified medium was 1.25 g obtained at 4.0 mg/l. Induced callus morphologically characterizes with whitish yellow colour and friable textured. Among these two growth regulators, Kn was found more efficient than BAP for callus proliferation with respect to days taken for callus initiation and induced callus weight. The results of current investigation give an idea of efficacy of BAP and Kn for morphogenetic effects on callus proliferation. Thus, for best economical use, the current study may help in identification of suitable growth regulator with appropriate level for getting maximum callus induction response of Aloe vera under in-vitro condition.
... e sexual reproduction of aloes is ineffective due to male sterility. us, they propagate primarily by means of sucker or offshoots [7,11,12]. Unfortunately, aloes' natural vegetative propagation is, likewise, very slow and ineffective to meet any demand of high-quality planting material for commercial cultivation [6]. A. trichosantha, belonging to the Tigrai and Eritrea floristic regions of the flora of Ethiopia and Eritrea, is one of the species being exploited for industrial purposes in Ethiopia. An Ethiopian company, specialized in producing jute bags for packing coffee, cereals, and oil seeds was granted to harvest A. debrana and A. trichosantha subsp. ...
This study was aimed to develop in vitro micropropagation protocol of Aloe trichosantha Berger using offshoots as explants. MS media supplemented with plant growth regulators helped explants develop shoots within about 14 to 17 days. The mean number of days to shooting has decreased from 16.8 ± 0.8 with 0.5/0.5 mg/L BAP/NAA supplement to 15.5 ± 0.5 with 2.0/0.5 mg/L BAP/NAA. While the mean shoot number has increased with increasing the concentration of BAP supplements, the reverse was true with mean shoot lengths, whereas supplement of 2.0/0.5 mg/L BAP/NAA has generated significantly more shoots (17 ± 3.8), and longer shoots were produced with the addition of 0.5/0.5 and 1.0/0.5 mg/L BAP/NAA. In regard to rooting, though higher concentrations of NAA have resulted in quick rooting, the rooting performance in terms of mean number and length of roots was better with low concentrations. All the plantlets subjected to greenhouse acclimatization in cocopeat have survived. Secondary acclimatization in composted and manured soil media has also resulted in 93 to 95% survival rate. Lighting conditions (nursery shade or direct sunlight) of secondary acclimatization did not lead to any difference in the survival rate of the plantlets.
... Nurashikin et al. (2010) obtained the greatest shoot regeneration (18.9 shoots/explant) from young shoot explants of Chlorophytum borivilianum from 3 mg L -1 BAP-supplemented MS medium. Choudhary et al. (2011) obtained the greatest shoot formation ratio of shoot tip explants of Aloe vera from BAP (1.0 mg L -1 ) + NAA (0.5 mg L -1 ) combination of MS medium. Razib et al. (2016) reported about 90% shoot regeneration from 2.0 mg L -1 BA + 0.5 mg L -1 NAA + 40.0 mg L -1 Ads (Adenine sulfide)-supplemented MS medium. ...
... In previous in vitro propagation studies conducted with different species, more successful bulblet and shoot regeneration were generally achieved in BA + NAA (Haibin and Jiajun, 2006;Yang et al., 2010;Sun and Jin, 2011;Zhang and Jia, 2014;Razib et al., 2016), BAP + NAA (Tang et al., 2009;Choudhary et al., 2011), only BAP, (Nurashikin et al., 2010;Jakhar et al., 2012;Kamaleswari et al., 2016;Pandey, 2016) or TDZcontaining (Çakmak et al., 2016;Kamaleswari et al., 2016) MS media. Generally, in the second and third set of my experiments, 0.5 mg L -1 BAP-containing MS media and TDZ + NAA combinations yielded more successful bulblet and shoot formations (Table 3 and 4). ...
Eremurus spectabilis M.Bieb is consumed as a vegetable because of its nutritious characteristics. The plants are also used for medicinal purposes, in the cut flower industry as an ornamental geophytes, and in industry as a natural adhesive. The aim of the present study was to improve the in vitro propagation protocol (in vitro germination and bulblet and shoot formation) of E. spectabilis. For this purpose, E. spectabilis seeds were in vitro germinated in four different nutrient media: Murashige and Skoog (MS), Gamborg (B5), White (WH), and Shenk and Hildebrandt (SH). To stimulate bulblet and/or shoot regeneration, hypocotyls of 35-40-day-old in vitro-germinated plantlets were cut into 0.5-1.0 cm pieces, and the resultant explants were cultured in MS media containing 2,4-dichlorophenoxyacetic acid (2,4-D) (0.5, 1.0, 2.0, and 4.0 mg L-1) + Kinetin (0.5 mg L-1), Thidiazuron (TDZ) (0.5, 1.0, 2.0, and 4.0 mg L-1) + 1-Naphthylacetic acid (NAA) (0, 0.1, 0.5, and 1.0 mg L-1), and 6-Benzylaminopurine (BAP) (0.5, 1.0, 2.0, and 4.0 mg L-1) + 2,4-dichlorophenoxyacetic acid (2,4-D) (0, 0.1, 0.5, and 1.0 mg L-1). The best outcomes for germination ratio (57.5%) were obtained from the B5 medium. In the third set of in vitro propagation experiments, 100% bulblet formation was achieved in TDZ (0.5 mg L-1) and NAA (0.5 and 0.1 mg L-1) combinations of MS media, and this was followed by 0.5 mg L-1 BAP-containing medium (81.3%). Shoot formation ratios with the same media combinations varied from 60-70%, and the number of shoots per explant varied from 1.4-2.4 shoots. Further in vitro propagation research is planned with larger bulb sizes to develop a protocol for rooting bulblets and/or shoots.
... Auxins, ABA, cytokinins, ethylene and gibberellins are commonly recognized as the five main classes of plant hormones. Auxins, cytokinins, and auxin-cytokinin interactions are usually considered to be the most important for regulating growth and organized development in plant tissue and organ cultures, as these two classes of hormones are generally required [9] . Auxins exert a strong influence over processes such as cell growth expansion, cell wall acidification, initiation of cell division, organization of meristems giving rise to either unorganized tissue (callus) or defined organs (generally roots) and promote vascular differentiation. ...
Gwarpatha (Aloe barbadensis Mill.) grown in warm tropical areas and is one of the most studied perennial shrubs worldwide. It has valuable medicinal properties and commercially used in pharmaceuticals, cosmetics, food industries and for many herbal preparations. It contains antioxidant vitamins which prevent damages caused by free radical, reducing risk of chronic diseases such as cancer, heart diseases and stroke. In nature Aloe barbadensis is propagated through lateral shoots which shows slow growth and are low in numbers and cultivation practice is expensive. For its large scale plantation there is a need to develop suitable and alternative method like in vitro micropropagation to meet the demand of farmers and pharmaceutical industries. Auxins, cytokinins and auxin-cytokinin interactions are considered to be the most important for regulating growth and organized development in tissue and organ cultures, as these two classes of hormones are generally required for in vitro propagation of gwarpatha. The present study aimed to develop collective information on protocol for rapid and high frequency in vitro propagation of Aloe barbadensis, looking through its increasing demand for pharmaceutical industry at global level.
... It propagates vegetatively, and its propagation rate is low [99]. Most parts of the plant are utilized by the drug and cosmetic industries [100,101]. It is generally used in the Ayurveda system to cure skin diseases. ...
The exploration of plant tissue culture implies aseptic development of explants in vitro in culture media comprising supplements and plant hormones and their hatching under controlled physical conditions (light, temperature and humidity). Tissue culture has been a subject of enthusiasm for quite a while, and lately it has turned into a valuable tool for agribusiness and medicines. Recently, tissue culture has developed as a science with a huge potential for large-scale plant production, particularly in horticulture and forestry. Quality planting materials of numerous species have been produced on a mass scale through tissue culture. The present chapter reviews the in vitro propagation studies of some commercially important plant species like Eucalyptus, Populus and Aloe.
... This is in contrast to the results found for tomato by Bhatia and Ashwath [15]. On the other hand, the results clearly showed that PVP improved the morphogenic capacity of meristems, which coincides with results for chestnut trees [16] and Aloe vera [17]. Phenolic oxidation is also a problem prevalent in the growth and propagation of established in vitro plants and has both an environmental and genetic component. ...
Background
Vegetative propagation of Fragaria sp. is traditionally carried out using stolons. This system of propagation, in addition to being slow, can spread plant diseases, particularly serious being viral. In vitro culture of meristems and the establishment of micropropagation protocols are important tools for solving these problems. In recent years, considerable effort has been made to develop in vitro propagation of the commercial strawberry in order to produce virus-free plants of high quality. These previous results can serve as the basis for developing in vitro-based propagation technologies in the less studied species Fragaria chiloensis.
Results
In this context, we studied the cultivation of meristems and establishment of a micropropagation protocol for F. chiloensis. The addition of polyvinylpyrrolidone (PVP) improved the meristem regeneration efficiency of F. chiloensis accessions. Similarly, the use of 6-benzylaminopurine (BAP) in the culture media increased the average rate of multiplication to 3–6 shoots per plant. In addition, the use of 6-benzylaminopurine (BAP), had low levels (near zero) of explant losses due to oxidation. However, plant height as well as number of leaves and roots were higher in media without growth regulators, with average values of 0.5 cm, 9 leaves and 4 roots per plant.
Conclusions
For the first time in Chilean strawberry, meristem culture demonstrated to be an efficient tool for eliminating virus from infected plants, giving the possibility to produce disease free propagation material. Also, the addition of PVP into the basal MS medium improved the efficiency of plant recovery from isolated meristems. Farmers can now access to high quality plant material produced by biotech tools which will improve their technological practices.
Electronic supplementary material
The online version of this article (doi:10.1186/s40659-017-0125-8) contains supplementary material, which is available to authorized users.
... Most of them (80 -90 %) are ornamental plants. Micropropagation techniques found wide applications with several succulents of the Agave (Robert et al., 1987), Aloes (Choudhary et al., 1987), Hylotelephium (Nakano et al., 2005), Sedum (Wojciechowicz, 2009), and Kalanchoe (Saifullah et al., 2006) genera. Each year this list is supplemented with new species. ...
Agaves are a very important group of plants. They are popular ornamentals but they are also used in the production of drugs, cosmetics, drinks, food and fodder. Unfortunately, due to the growing influence of anthropopressure, some of them are threaten with extinction. Therefore, in order to always be able to meet the growing demands of the market, novel biotechnolog-ical tools need to be applied in the production of these species. Micropropagation, i.e. vegetative multiplication of plants under aseptic, strictly controlled conditions and with the use of syn-thetic media, is the most commonly applied aspect of plant tissue cultures. The technique reduces time, space and costs required for the production of plants. Over time, several micropropaga-tion techniques have been developed also with agaves. The aim of the present review is to present the current achievements and problems associated with micropropagation of the most impor-tant agave species.
