ABSTRACT:In vitro selected salt tolerant saplings of Morus alba (cv. Sujanpuri) were raised from nodal explants with axillary buds collected during three different periods of the year. The growth and shoot/root multiplication of the nodal explants collected between November to February and July to October were found to be better than those collected between March to June. In cultures, shoot multiplication was induced by the application of 2.5 mg/l of 6-benzylaminopurine (BAP) and 0.3 mg/l of gibberellic acid (GA 3 ), while rooting by 1.0 mg/l of indolebutyric acid (IBA). Sodium chloride (NaCl) was added to induce salt stress and its concentration was gradually increased from 0.1% (w/v) onwards. The salt tolerance was observed up to 0.4% (w/v) NaCl and 100% mortality of explants was noted above this concentration. The inclusion of Arbuscular Mycorrhizal (AM) fungi and Azotobacter chroococcum to tissue culture of raised saplings during acclimatization enhanced their survival and resulted in a significant increase of plant growth. After the transfer of plants to salt affected wasteland, only NaCl-treated saplings survived, whereas those developed without NaCl resulted in 100% mortality. plying salt (NaCl) stress and if exploiting AM fungi and Azotobacter during hardening could enhance the survival percentage of developed plants on salt affected land. Keywords: in vitro; MATERIALS AND METHODSExplant collection: Growing shoots of Morus alba L. cv. Sujanpuri were collected from one year old mulberry plants cultivated on normal soil (pH = 7.5, EC = 0.110 mmhos/cm) at Micromodel, IIT, Delhi during the following months: A) March to June, B) July to October, C) November to February. Nodal segments with one axillary bud each (1.5 to 2.0 cm) were excised, washed and disinfected first with 0.7% (w/v) bleach solution (sodium hypochloride) for 10 min and with 0.1% aqueous mercuric chloride solution for next 10 minutes and then rinsed 5 to 6 times with sterile distilled water.Shoot multiplication: The Murashige and Skoog's (MS) (1962) medium containing 3% (w/v) sucrose, 2.5 mg/l of BAP and 0.3 mg/l of GA 3 with 1.0% agar and pH 5.8 was used as shoot multiplication medium. For induction of salinity stress, the medium was supplemented with 0.1% NaCl. The surface sterilized explants were inoculated vertically onto the culture medium with one explant per test tube and twenty test tubes were kept for each treatment.Cultures were grown under cool white fluorescent tubes with irradiance of 24 µmol m 2 /s, 16 h photoperiod and at the temperature of 25 ± 1°C. After multiple shoot induction, the NaCl concentration of medium was gradually increased from 0.1 to 0.4% till 100% mortality of explants observed and average shoot length and number of shoots per explant was noted.Root induction: For rhizogenesis, well developed shoots (5.0 cm long) with 2-3 leaves were excised from cultures and cultured on MS supplemented with 1.0 mg/l of IBA, 3.0% sucrose and NaCl (from 0.1 to 0.4%). For root production, shoots were inoculated on medium with the...
In vitro tissue culture of hard woody, endangered, medicinal plant Coscinium fenestratum is most challenging to plant tissue culturists. In the present study, petiole and leaf explants of Coscinium fenestratum were induced to form callus when cultured on vermicompost extract media along with coelomic fluid. Suspension medium was developed using vermicompost extract and coelomic fluid in 3:1 ratio. Phytochemical analysis of the alkaloid berberine was confirmed from callus, suspension cell culture and suspension medium by Thin Layer Chromatography and High Performance Liquid Chromatography. Vermicompost and its extracts with coelomic fluid have shown maximum (100 per cent) response of callus induction. Callus mass enlarged with increasing concentration of coelomic fluid and callus growth was assessed from the biomass. Incubation of culture tubes in dark supported callus development significantly. The Rf value of 0.36 confirmed the presence of berberine by Thin Layer Chromatography. Qualitative analysis confirmed the presence of alkaloid berberine with the retention time of 2.8 minutes similar to that of standard reference sample from Sigma chemicals, USA. The suspension medium turned deep yellow because of the release of the alkaloid. Vermicompost and its extracts along with coelomic fluid have shown the economical approach for micropropagation of economically and medicinally important plants. S. Kashyap et al.
Humus is a complex material formed during the breakdown of organic matter. Earthworm castings contain a high percentage of humus. These humic substances are capable of improving plant nutrition and growth, reminiscent of hormones. Residue obtained after acid-base treatment of vermicompost was used as plant tissue culture media for the micropropagation of Bacopa monnieri. Tukey's Studentized Range (HSD) Test has clearly indicated that development of leaves from nodes was significantly higher in humin alone without any supplements. The F value was 5.5 and Pr > F was 0.0087. The weight of the plantlets (in milligrams) was significantly higher in the humin medium and humin supplemented with vitamins and micronutrients but was least in the humin medium with growth regulators. The difference observed was at the level of < 0. 0001. The explant development on humin alone has shown the significant percentage survival of in vitro micropropagated plantlets when compared with other treatments. Explants responded maximum to humin only but not so on adding the supplements and the level of significance was at Pr > F was < 0.0001. The maximum growth stimulatory effect was found in aqueous extract of vermicompost which can be made out from the Student's t-test performed to compare the means of weight of plantlets grown on humin and vermicompost extract media with and without chemical supplements respectively. The probability of this result, assuming the null hypothesis is 0.0014.
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