Effect of arbuscular mycorrhizal fungi on the acclimatization of micropropagated banana plantlets

 The effect of arbuscular mycorrhizal fungi (AMF) on micropropagated banana plantlets was evaluated during the acclimatization period. Plants were inoculated with Acaulospora scrobiculata, Glomus clarum or Glomus etunicatum. After cultivation in a greenhouse for 3 months, height, leaf area, fresh weight and dry matter of root and shoots, level of AMF colonization, nutrient level, photosynthesis and transpiration rate, water potential and stomatal conductance were measured. The number of AMF spores produced in each treatment was also determined. Plantlets inoculated with AMF had greater height, leaf area and fresh weight of shoots and roots, as well as higher rates of photosynthesis and transpiration than controls. Plants inoculated with Glomus were superior in most of the evaluated parameters. Accepted: 24 May 1999     

Efficient In vitro Plant Regeneration Protocol from Leaf Explant of Jatropha curcas L — A Promising Biofuel Plant

An efficient in vitro plant regeneration from leaf-disc culture of Jatropha curcas L has been established. Adventitious shoot buds along with callus were induced from leaves of 2-year-old J. curcas plants cultured on Murashige and Skoog’s (MS) medium supplemented with TDZ (2 μM) BAP (2 μM) and IBA (1 μM), wherein 63.3% leaf explants responded. The multiplication of shoots was achieved from the adventitious shoot buds after transferring them to shoot induction medium. The highest number of shoots (9.7/explant) was achieved after 6 weeks of culture on MS medium containing 3 μM of BAR The welldeveloped shoots were rooted on MS medium supplemented with IBA (1.5 μM) with the rooting frequency of 53.3%. Addition of phloroglucinol (200 μM) to the medium enhanced the frequency of rooting to 76.7%. Regenerated plantlets were successfully transferred to field after initial acclimatization.     

Efficient Plant Regeneration via Shoot Tip Explant in Jatropha curcas L

An efficient regeneration method via shoot tip explant has been developed for Jatropha curcas, which is a medicinally as well as economically important plant. Shoot tips were proliferated on MS medium incorporated with BAP (2.0 mg l^?1) and IAA (0.5 mg l^?1) along with adenine sulphate, glutamine and activated charcoal. In vitro produced shoots were induced to root on IBA (0.5–5.0 mg l^?1) added to half strength MS medium. The highest frequency of root induction was on the medium with 3.0 mg l^?1 IBA. Regenerated plantlets were successfully transferred to field after initial acclimatization.     

High-Frequency Regeneration by Abscisic Acid (ABA) from Petiole Callus of Jatropha curcas

Jatropha curcas L. is attaining worldwide interest as an important biofuel crop. Experiments were conducted to improve the prevailing micropropagation technique as well as to develop a new ex vitro rooting method for J. curcas plant regeneration. Regeneration and ex vitro rooting efficiency was enhanced by augmenting the culture medium with abscisic acid (ABA). Different concentrations of 6-benzylaminopurine (BAP) and indole-3-butyric acid (IBA) were tested for callus generation from both in vitro and in vivo explants (leaf and petiole) on Murashige and Skoog (MS) medium. The best regenerative callus was achieved on MS medium supplemented with BAP (4.44 μM) and IBA (2.45 μM) from in vitro-cultured petioles. Highest regeneration (91%) was achieved by culturing petiole callus on MS medium supplemented with BAP (8.88 μM), IBA (0.49 μM), and ABA (1.9 μM), whereas 61% regeneration was obtained from in vitro leaf callus. Shoot proliferation and elongation was achieved on BAP (2.22 μM) and IAA (8.56 μM) with 10–13 shoots per explants. Highest rooting (65%) was achieved from M1 shoots (BAP, IAA, and ABA) on MS medium supplemented with IBA (2.45 μM), naphthaleneacetic acid NAA (0.54 μM), and 0.02% activated charcoal. Ex vitro rooting of 1-mo-old M1 shoots obtained from the charcoal-containing medium resulted optimum rooting (>72%) when transferred to polybags containing sterile sand. The plantlets were successfully acclimatized in soil with more than 98% survival rate in the greenhouse.     

Assessment of Genetic Stability in Three Generations of In Vitro Propagated <Emphasis Type="Italic">Jatropha curcas</Emphasis> L. Plantlets Using ISSR Markers

Tissue culture has been widely employed in Jatropha curcas L. for the clonal multiplication of superior genotypes. However, the evaluation of genetic stability is necessary to detect somaclonal variants. In this context, the present aim was to evaluate the genetic stability of J. curcas plantlets, obtained via indirect organogenesis, by means of ISSR markers. To supply the explant sources for in vitro propagation, the first generation of plants was produced from in vitro germination of J. curcas seeds. Fragments of cotyledonary leaves were inoculated into medium supplemented with 1.5 mg L^?1 BAP and 0.05 mg L^?1 of IBA for induction of callogenesis. The resulting calli were transferred to bud induction medium. Subsequently, the buds were cultured in medium for elongation, giving rise to the second generation of plants. These plants provided new buds, which were excised and subcultured in elongation medium, yielding a third generation of plants. To evaluate genetic stability in three plant generations, twelve ISSR primers were used, resulting in 124 bands showing 41.93 % of polymorphism. Increase was observed in the level of somaclonal variation (SV) over the generations. The present study reports, for the first time, the analysis of genetic stability in J. curcas plantlets regenerated via indirect organogenesis by means of ISSR markers. The results suggest that the indirect route is associated to higher levels of genetic instability, which also increased with successive subcultures. The ISSR markers were efficient in detecting SV, and the generated genetic variability may be useful for breeding programs.     

Improved in vitro rooting and hyperhydricity in regenerating tissues of Thapsia garganica L.

Stock cultures of Thapsia garganica grown on Murashige and Skoog agar medium (1962) (MS) (0.8% agar [w/v]; pH 5.8) with 0.5 mg l⁻¹ NAA and 1.5 mg l⁻¹ BA were best rooted by subjecting to half strength MS liquid medium with IBA (10 mg l⁻¹) for 3 days prior to transfer to medium without plant growth regulators. A rooting frequency of 60% was noted with seven roots per rooted plant. Rooting was apparent after 10 days. The present study also aimed at reducing the occurrence of hyperhydric plants. The inclusion of 2% polyethylene glycol (w/v) in the growth medium or ventilation of cultures prior to acclimatization resulted in the production of plants true to type, closely resembling wild plants. Those plantlets that had been rooted and exposed to a better vented environment were able to acclimatize readily. Tissue culture propagation is therefore beneficial to the conservation of the medicinally important species, Thapsia garganica.     

Exogenous systemin has a contrasting effect on disease resistance in mycorrhizal tomato (<Emphasis Type="Italic">Solanum lycopersicum</Emphasis>) plants infected with necrotrophic or hemibiotrophic pathogens

A study was performed to determine the effect of the systemin polypeptide on the bio-protective effect of arbuscular mycorrhizal fungi (AMF) in tomato plants infected with Alternaria solani, Phytophthora infestans or P. parasitica. Before infection, tomato plants were colonized with two different AMF, Glomus fasciculatum or G. clarum. In addition, a group of inoculated plants was treated with systemin, just after emergence. The exogenous application of systemin marginally suppressed the resistance against A. solani leaf blight observed in G. fasciculatum mycorrhizal plants but significantly enhanced it in plants colonized with G. clarum. Systemin induced resistance to P. parasitica in leaves of G. fasciculatum mycorrhizal plants, in which AMF colonization alone was shown to have no protective effect. Conversely, none of the treatments led to resistance to root or stem rots caused by P. infestans or P. parasitica. The above effects did not correlate with changes in the activity levels of β-1,3-glucanase (BG), chitinase (CHI), peroxidase (PRX), and phenylalanine ammonium lyase (PAL) in leaves of infected plants. However, they corroborated previous reports showing that colonization by AMF can lead to a systemic resistance response against A. solani. Systemic resistance to A. solani was similarly observed in non-mycorrhizal systemin-treated plants, which, in contrast, showed increased susceptibility to P. infestans and P. parasitica. The results indicated that the pattern of systemic disease resistance conferred by mycorrhizal colonization was dependent on the AMF employed and could be altered by the exogenous application of systemin, by means of a still undefined mechanism.     

Micropropagation protocol for Antigonon leptopus an important ornamental and medicinal plant

The effect of some factors on in vitro consecutive micropropagation behavior of Antigonon leptopus was examined including those of culture establishment, shootlets multiplication, rooting and acclimatization stages. The highest percent of aseptic cultures and survival of explants (100%) were obtained as a result of using Clorox 10% for 3?min followed by MC 0.1% for 2?min while, using each of them individually (Clorox 20% or MC 0.1%) for 5?min caused the highest percent of shoot formation. During the multiplication stage, the highest percent of shoot formation was reached to 100% with repeating culture of explants (two times) on MS medium supplemented with 2ip at 1.0 and IBA at 0.2?mg/l. The highest numbers of shootlets/explant were obtained when 2.0?mg/l of BAP or 0.5?mg/l BA?+?0.2?mg/l of IBA were added to MS culture medium. Culturing the explants on MS medium supplemented with 2ip at 0.5 or 1.0?mg/l each combined with 0.2?mg/l of IBA showed the longest shootlets. Reducing the strength of culture media to ½ or ¾ had promotion effect on rooting formation of shootlets. The best results of plant acclimatization (survival percent, plant height and root length) were obtained by using sand or peat moss soil. The amplified DNA fragments using B7, B9 and C19 primers for mother and micropropagated plants showed that the produced pattern by primer B7 had a maximum number of 10 bands of DNA fragments with molecular size ranging between 1025.57 and 176.36?bp, micropropagated plants showed 95.2% similarity in relation to mother plant.     

Effect of the ectomycorrhizal fungus Hebeloma cylindrosporum on in vitro rooting of micropropagated cuttings of arbuscular mycorrhiza-forming Prunus avium and Prunus cerasus

 The effect of different genotypes of the ectomycorrhizal fungus Hebeloma cylindrosporum on in vitro rooting of micropropagated cuttings of Prunus avium and P. cerasus was studied in an attempt to determine whether ectomycorrhizal fungi could enhance in vitro adventitious root formation in plants which form arbuscular endomycorrhizas. The rooting percentage of P. avium cuttings was approximately 16% in the absence of hormonal treatment; it increased up to 30% in the presence of 5.7 μM IAA which was the most favourable auxin concentration. The rooting percentage of cuttings cultivated in the absence of IAA was enhanced by all the studied strains of H. cylindrosporum. It ranged from 50 to 60% with the IAA-overproducing mutant D 111 or the wild-type dikaryon D1, to 100% in the presence of the mutants 331 or D 117. The cuttings of P. cerasus showed a higher rooting ability than those of P. avium since approximately 40% of them were able to root in the absence of hormonal treatment. Except for the mutant D117, their rooting percentage was not significantly improved by H. cylindrosporum. Fungal inoculation also affected the survival of cuttings at acclimatization: 50% of the uninoculated P. avium cuttings survived whereas the survival percentage of inoculated cuttings ranged from 30 to 100% depending on the fungal genotype. With P. cerasus, the percentage of survival of uninoculated cuttings ranged from 85 to 100% and fungi either did not significantly improve it or lowered it. At acclimatization fungal hyphae could be observed in close contact with adventitious roots, but they did not establish mycorrhizal association. The shoot height of P. avium plantlets obtained from inoculated cuttings was not significantly different from that of plantlets originating from uninoculated ones. By contrast, fungal inoculation generally depressed the growth of acclimatized P. cerasus plantlets. The possibility of using ectomycorrhizal fungi as a tool to enhance rooting of micropropagated cuttings of plants which do not form ectomycorrhizas is discussed. Received: 25 November 1996 / Accepted: 2 June 1997     

Tissue Culture Propagation of Elite Plant of Aloe vera Linn

Micropropagation protocol for an elite selection of Aloe vera syn A. barbadensis through enhanced axillary branching was standardized. Murashige and Skoog medium containing 1 mg l^?1 BA and 0.2 mg l^-1 IBA gave highest multiplication. Citric acid at 10mg l^-1 and liquid medium improved the shoot multiplication. Hundred per cent microshoots produced rooted plantlets within 15 days of culture on hormone-free agar medium. Liquid medium during rooting stage decreased the number of shoots showing rooting response. The plants were successfully transferred in the soil and were morphologically similar to mother plants.     

GIS-facilitated in vitro propagation and ex situ conservation of Achillea occulta

A Geographical Information Systems (GIS)-facilitated approach for the in vitro propagation and ex situ conservation of the conservation priority species Achillea occulta is presented. To realize the species?? ecological requirements, the coordinates of the original habitat were linked with thematic layers derived from digital databases in a GIS environment. From wild plants, shoot tips were established in vitro in a basal medium with 4???M 6-benzyladenine (BA) and 0.5???M indole-3-butyric acid (IBA). A modified basal MS medium (modMS, double amount of Fe) proved to be the most effective for in vitro adventitious shoot production. The effect of BA in combination with ??-naphthaleneacetic acid (NAA) or IBA on shoot proliferation was also tested. The highest number of new microshoots/explant (3.5), with 0.93?cm shoot height was obtained when the modMS was supplemented with 5???M BA and 2.5???M IBA. To evaluate the root induction ability, microshoots produced were transferred to modMS media supplemented with 0?C20???M IBA and 0?C20???M NAA. Rooting proved to be very difficult and only by adding 20???M IBA, a 12.5% rooting percentage was achieved. Acclimatization succeeded only during early spring. Young plants transplanted at the Balkan Botanic Garden of Kroussia produced flowers and seeds in the first year. This GIS-approach provided useful guidelines for A. occulta??s (a) effective propagation (selection of greenhouse temperatures, temperatures during in vitro culture, suitable period for cuttings and acclimatization of plantlets), and (b) ex situ cultivation (selection of watering regime, temperatures, locations and exposures for growing sites).     

Correction to: Performance of cowpea as influenced by native strain of rhizobia,lime and phosphorus in Samaru,Nigeria

Garden thyme (Thymus vulgaris L., Lamiaceae) is an important aromatic herb used for its medicinal values including antioxidant and antimicrobial properties. The present study was performed to analyze the changes in natural antioxidants after inoculation of in vitro propagated garden thyme plants with arbuscular mycorrhizal fungi (AMF). An efficient and low-cost protocol for large-scale multiplication of this aromatic plant was developed. The explants were cultured on full and half strength Murashige and Skoog (MS) medium containing indole-3-butyric acid (IBA). The maximum number of shoots and roots was obtained on ½ MS medium supplemented with 0.1 mg L^?1 IBA after 4 weeks of culture. The successfully adapted in vitro plants (survival rate 95%) were inoculated with arbuscular mycorrhizal fungi (Claroideoglomus claroideum, ref. EEZ 54). Plants were then transferred into field conditions. Mycorrhizal fungi enhanced the activity of some soil enzymes, acid and alkaline phosphatase, urease as well as the levels of extractable glomalin-related proteins in plant rhizosphere. Arbuscular mycorrhizal associations with higher plants promote the accumulation of antioxidant metabolites such as phenols and flavonoids and increase the activity of antioxidant enzymes. The results from the present study suggest enhanced antioxidant capacity of the inoculated T. vulgaris plants which was due mainly to increased accumulation of phenolic compounds (total phenols and flavonoids) together with stimulation of the activity of superoxide dismutase (SOD) and guaiacol peroxidase (GPO).     

In vitro plant regeneration from cotyledon fragments of the olive tree (Olea europaea L)

Olive tree (Olea europaea L) plantlets were regenerated from cotyledon segment calli on a modified olive medium (OMc) supplemented with 2iP alone or in combination with indol-3-butyric acid (IBA). Cell division in the explants was initially induced on OMc medium with high auxin (5 mg·l⁻¹ of IBA) and low cytokinin (0.2–0.5 mg·l⁻¹ of 2-isopentenyladenine (2iP) or zeatin riboside) content. Calli were then transferred to the same medium with different levels of IBA and/or 2iP in order to promote further development and obtain calli bearing either roots or shoots. On OMc medium, 1 mg·l⁻¹ of IBA induced the maximum of rooting, while shoot induction was greater when the medium was supplemented with 4 mg·l⁻¹ of 2iP. Shoot induction mainly occurred from calli of cotyledon fragments proximal to the embryo axes. Whole plantlets were obtained when the regenerated shoots were stimulated to produce adventitious roots on OMr medium with 1 mg·l⁻¹ of IBA or naphthaleneacetic acid (NAA). After root elongation on OMe medium without auxin, plantlets were transfered to peat and soil conditions where about 75–80% were able to survive. A certain variability was detected between regenerated olive plants.     

Mycorrhizal fungi enhancement of growth and gas exchange of micropropagated guava plantlets (Psidium guajava L.) during ex vitro acclimatization and plant establishment

Psidium guajava L.) plantlets was determined during acclimatization and plant establishment. Guava plantlets were asexually propagated through tissue culture and grown in a glasshouse for 18 weeks. Half of the plantlets were inoculated with a mixed endomycorrhiza isolate from Mexico, ZAC-19, containing Glomus diaphanum, G. albidum and G. claroides. Plantlets were fertilized with modified Long Ashton nutrient solution that supplied 11 μg P ml⁻¹. Gas exchange measurements were taken at 2, 4, 8, and 18 weeks after inoculation using a portable photosynthesis system. All micropropagated guava plantlets survived transplant shock. After 6 weeks, mycorrhizal plantlets had greater shoot growth rates and leaf production than non-mycorrhizal plantlets. This also corresponded with increased photosynthetic rates and stomatal conductance of mycorrhizal plants. By 18 weeks, mycorrhizal plantlets had greater shoot length, leaf area, leaf, stem, and root dry mass. However, gas exchange was comparable among treatments, in part because the container size was restricting growth of the larger mycorrhizal plantlets. Non-mycorrhizal plantlets had greater leaf area ratios and specific leaf areas than mycorrhizal plantlets. Increased leaf tissue mineral levels of P, Mg, Cu, and Mo also occurred with mycorrhizal plantlets. Roots of inoculated guava plantlets were heavily colonized with arbuscules, vesicles and endospores. Guava plantlets were highly mycotrophic with a mycorrhizal dependency index of 103%. Accepted: 27 December 1999     

In vitro propagation of a semi-dwarfing cherry rootstock

A successful in vitro propagation system for the semi-dwarfing cherry rootstock Maxma-14 (Prunus avium L.) has been developed. Shoot tips and axillary buds were successfully established in vitro. Multiplication rate of about 6 was achieved over a 4-week period using Murashige and Skoog medium with 4.44 μM benzyladenine and 0.49 μM indole-3-butyric acid (IBA). Rooting occurred within 4 weeks on liquid and agar-gelled media containing 0.49 μM NAA or 0.49, 2.45 μM IBA. On liquid media, a maximum rooting efficiency of up to 100% was obtained. However, high concentrations of auxins delayed the time of root initiation for 3–5 days. Acclimatization was affected directly by rooting conditions. Survival was best when plantlets were transferred to pots after a short period of root emergence on rooting media. Multiplication medium was also important for successful acclimatization. Shoots transferred to rooting media from that with kinetin resulted in better acclimatization and survival than that derived from media with benzyladenine. Further, plantlets rooted on liquid media had better survival than that rooted on agar-gelled media. This revised version was published online in June 2006 with corrections to the Cover Date.     

Micropropagation of Chokecherry and Pincherry cultivars

In vitro culture establishment, shoot proliferation and ex vitro rooting responses of chokecherry (Prunus virginiana L.), `Garrington', and pincherry (P. pensylvanica L.f), `Mary Liss' and `Jumping Pound', were examined using various combinations of growth regulators. Dormant winter buds were used as explants. MSMO medium supplemented with 0.49 μM IBA and either 4.44 or 8.87 μM BA was found to be optimal for culture initiation of both species and cultivars. GA₃ (28.89 μM) significantly reduced (p=0.0001) the number of successfully established cultures. BA concentrations 8.87–12.82 μM gave optimal shoot proliferation in chokecherry and 4.44 μM BA in both cultivars of pincherry. Auxin treatments were required for ex vitro rooting of approximately 10 mm long shoots in peat/perlite (1:1 v/v) mixture, at 25 °C, under mist. The best rooting (84%) was obtained with IBA/NAA (9.80/2.69 μM). A commercial rooting powder, Rootone F, containing IBA/NAA (0.057/0.067%) mixture, was also effective (75%). The ex vitro rooted plantlets did not require any additional acclimatization prior to transplanting to the regular greenhouse conditions. This revised version was published online in August 2006 with corrections to the Cover Date.     

Changes in activity of antioxidant enzymes and photosynthetic machinery during acclimatization of micropropagated Cassia alata L. plantlets

The stimulatory effect of thidiazuron (TDZ) has been investigated in shoot multiplication for a simple, efficient, rapid, and commercially applicable regeneration protocol of an important medicinal plant, Cassia alata. Furthermore, the effects of an increased photosynthetic photon flux density (PPFD) on photosynthesis, the functioning of the photosynthetic apparatus, and the response of the antioxidant enzymatic system were studied during the ex vitro establishment of micropropagated plantlets. Multiple shoots were induced by culturing nodal explants excised from an aseptic seedling on Murashige and Skoog (MS) medium supplemented with various concentrations (1.0, 2.5, 5.0, 7.5, or 10.0 μM) of TDZ for different treatment durations (2, 3, 4, or 6 wk). The highest number of shoots (17.9?±?0.3) and longest shoot length (4.6?±?0.1 cm) were achieved when explants were exposed to 5.0 μM TDZ for 4 wk and subsequently subcultured on growth regulator-free MS medium for 8 wk. In vitro rooting of isolated shoots was best achieved on full-strength MS medium containing 0.5 μM indole-3-butyric acid (IBA). The micropropagated shoots with well-developed roots were successfully established in pots containing Soilrite? followed by garden soil and grown in greenhouse with an 85% survival rate. During the acclimatization period, significant changes in the activity of the antioxidant enzymatic system were observed. An increase in superoxide dismutase (SOD) activity was measured throughout the acclimatization period. Likewise an upregulation of catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) enzyme activities were also observed. Pigment (chlorophyll a and b and carotenoids) content in ex vitro-formed leaves was significantly higher compared with those grown in vitro. These observed changes reflected the ability of plants to develop an antioxidant enzymatic defense system aiding in survival against oxidative stress and in reducing release of free radicals.     

In Vitro Method for Propagation of Centella asiatica (L) Urban by Shoot Tip Culture

A rapid clonal propagation system has been developed for the medicinally important herb Centella asiatica (L) Urban by shoot tip (2–3 cm long) culture. The shoot tips isolated from mature plants were inoculated on MS medium incorporated with BA alone or in combination with NAA and Kn. The optimum number of shoots (3.38) with optimum number of leaves per shoot (4.25) were attained on MS medium supplemented with 4.0 mg l^?1 BA and 0.1 mg l^?1 NAA. On transferring the microshoots on full strength MS medium supplemented with various concentrations of IBA (1.0-3.0 mg l^?1) and NAA (0.5-2.0 mg l^?1), profuse rooting (46.8 per shoot) was obtained in MS basal medium with 2.0 mg l^?1 IBA with root length of 19.7 cm. Well rooted plantlets were acclimatized successfully by adjusting the temperature and humidity for 3–4 weeks after transfer to pots filled with sterilized vermiculite soil: sand (1:1)mixture. This micropropgation protocol could be useful for raising a stock of genetically homogenous material for field cultivation within a very short period.     

Vesicular arbuscular mycorrhizal fungi improves establishment of micropropagated Leucaena leucocephala plantlets

Investigations were carried out to achieve cent per cent transplantation success of micropropagated Leucaena leucocephala (a fast growing multipurpose leguminous tree species) plantlets using two vesicular arbuscular mycorrhizal fungi, Glomus fasciculatum and Glomus macrocarpum. Plantlets were obtained by rooting the shoots [obtained through; hypocotyl callus in presence of 10^-5M BAP + 10^-6M NAA; and axillary bud sprouting from cotyledonary and other nodes in presence of 10^-5M BAP, on Gamborg's B5 medium], on half strength B5 medium supplemented with 5×10^-6M IBA. Subsequent to the nodulation of their roots with Rhizobium (strain PRGL 001)in soilrite, these plantlets were tranferred to sterilized garden soil by laying inoculum of either Glomus fasciculatum or Glomus macrocarpum around their roots. Only 20% of the plantlets survived in soils lacking VAM fungus. In contrast, cent per cent of the plantlets of Leucaena leucocephala established very well and showed good growth in VAM inoculated soil. Roots of the later plantlets showed presence of both external and internal hyphae with well formed arbuscules and vesicles confirming the establishment of good mycorrhizal association. These studies convincingly demonstrate that the mycorrhizal association help in successful establishment of tissue culture raised plantlets of Leucaena leucocephala in the field conditions by alleviating the transplantation shock. This revised version was published online in June 2006 with corrections to the Cover Date.     

In vitro regeneration in Sarcostemma acidum (Roxb.) –an important medicinal plant of semi-arid ecosystem of Rajasthan, India

An efficient regeneration protocol for Sarcostemma acidum – an important medicinal plant has been established. Callus initiated from nodal explant on MS medium with 2.0 mg?L^?1 of NAA + additives. Callus initiated was subcultured on MS medium containing various concentrations of NAA or 2,4-D. Out of these combinations, MS medium +1.0 mg?L^?1 of NAA + additives was found to be effective for the multiplication of callus. Subculture was done after an interval of 20–22 days. For differentiation of callus BAP or Kinetin alone was found to be less effective. Maximum frequency of shoot regeneration recorded on MS medium +1.0 mg?L^?1 of BAP?+?0.5 mg?L^?1 of Kinetin and 0.1 mg?L^?1 of NAA + additives. The in vitro differentiated shoots were excised and inoculated on 1/4 strength MS medium +2.0 mg?L^?1 of IBA?+?0.02 % activated charcoal for in vitro rooting. Maximum response (90 %) was recorded on this medium. In vitro differentiated shoots were inoculated on autoclaved soilrite® after treatment with root inducing auxins. Ex vitro rooting in this plant species has been reported for the first time. Eighty five percent of the shoots rooted under ex vitro conditions. Both in vitro and ex vitro rooted plantlets were hardened in a green house.