Browsing by Author "Mini Balachandran"
Now showing 1 - 2 of 2
Results Per Page
Sort Options
ThesisItem Open Access Improvement of Anthurium andreanum Lind. in Vitro(Department of Pomology and Floriculture, College of Horticulture, Vellanikkara, 1998) Mini Balachandran; KAU; Ramachandran Nair, SFor refining and establishing micropropagation system for Anthurium andreanum cv. Dragon's Tongue, studies were carried out, to solve many of the problems faced in the micropropagation of the variety, such as, improper foliage development, prolonged period for callus induction and multiplication. Investigations were conducted also to explore alternative explants for micropropagation, induction of somatic embryogenesis and development of artificial seeds, crop improvement through induction of somaclonal variation and radiation breeding in vitro. The study was carried out during 1995-98 at the Plant Tissue Culture Laboratory, Kerala Horticulture Development Programme (R & D), Kerala Agricultural University, Vellanikkara. Surface disinfestation with 70 per cent ethyl alcohol wipe followed by treatment with 0.1 per cent HgCl2 for eight minutes recorded maximum survival (99.74%) of cultures in the case of leaf explants. For spadix explants, ethyl alcohol wipe (70%) + emisan (0.1 %) dip for three minutes followed by HgCI2 (0.1 %) for 10 minutes recorded maximum survival (87.56%) of the cultures. , Among the different explants tried (in vivo and in vitro derived explants) callus induction was maximum in the case of in vivo leaf explants. Callus was initiated (86.50%) within 51 days compared to 90 days reported earlier, when cultured In darkness on to the culture media, Nitsch-white (NW) + kinetin 0.5mgl-1 + 2.4-0 0.3 rngl-1 + sucrose 20 gl-1+ glucose 10 gl-1 + agar 6.0 gl-1. Spadix explants and in vitro derived explants (leaves, nodes, petiole and roots) showed good response to callus induction treatments. Among the in vitro derived explants, root explants recorded the maximum callus multiplication. Callus induction from spadix explants was better (59.85%) in half strength MS basal medium supplemented with 2,4-0 0.3 mgl-1 + kinetin 0.5 mgl-1 +Sucrose 30 gl-1 + agar 6 gl-1. Maximum shoot regeneration (92%) was observed after 46 days in Nitsch media supplemented with BAP 0.5 mgl-1 in the case of leaf callus. In the case of callus derived from spadix explants, half strength MS media supplemented with BAP 2.0 mgl-1 + kinetin 2.0 mgl-1 recorded maximum response (71.42). Multiplication rate (22 per culture vessel) of the shoots and growth and development of the leaves and shoots were better in conical flasks (200ml size). For quicker establishment of the plantlets, mud pots gave the best results (85.33%). Incorporation of vesicular -arbuscular mycorrhizae (VAM) Glomus so., into the potting mix improved the growth of the plantlets ex vitro. Among the explants tried for somatic embryogenesis, in vitro derived leaves (53%) and petiole (18.90%) and immature seeds (8.33%) showed positive response. Induction of somatic embryoids was observed in the media, Nitsch-White (NW) supplemented with 2,4-0 1.5 mgl-1 + kinetin 0.15 mgl-1 + sucrose 20 gl-1 + glucose 10 gl-1 + glutamine 200 mgl-1 + agar 6 gl-1 in explants derived from in vitro leaves and petiole. For immature seeds, response was observed in Nitsch media supplemented with 2,4-02.0 mgl-1 + kinetin 0.3 mgl-1 + sucrose 20 gl-1 + glucose10 gl-1 + glutamine 200 mgl-1. Germination of the somatic embryoids was highest in half strength MS media supplemented with BAP 0.1 mgl-1 + glutamine 200 mgl-1 . Viability of the somatic embryoids was observed to be very low (5-10 days). Encapsulation of somatic embryoids was achieved with calcium chloride at 50 ~M and sodium alginate at 3 per cent level • "j After encapsulation, somatic embryoids can be stored up to 20 days without much loss in capacity for germination (15.50%). Germination per cent of encapsulated somatic embryos was improved (39%) after a low temperature storage (4 0C) for 20 days. Rudimentary leaves were observed in plantlets regenerated from ninth and tenth subculture. The colour of the leaves in such plantlets was observed to be pale green. After transplanting also, the plants exhibited poor leaf growth. In such plants the leaf area remained smaller than other plants. But, chlorophyll development was normal. For radiation breeding using y-rays, the irradiation doses above 150 Gy were found to be lethal to callus as well as for shoot tips. Maximum response in terms of plant height, plant spread and leaf area was recorded at lower dose of 50 Gy. Most significant variation observed in in vitro regenerated plantlets compared to mother plants was the reduction in leaf area of mutants regenerated from explants irradiated with 150 Gy. Height of the plant was also less at higher doses of ƛ-irradiation. The plants obtained from the trial on induction of somaclonal variation and radiation breeding were screened for possible mutants and somacional variants using morphological characters, biochemical markers and cytological technique. Morphological characters were recorded for observing possible variability, at periodical intervals after transplanting. Dwarf mutants were observed at higher doses of y-irradiation. The biochemical studies using isozymes revealed no difference among the plants regenerated from different subcultures and different doses of irradiation. But, difference was observed between the non-irradiated and the irradiated plants for the number of bands produced. Peroxidase isoenzyme was found to be the most stable and was expressed in plants regenerated from different subcultures as well as those from irradiated cultures. Five bands were resolved in the case of plants from the repeated subculturing and two bands were resolved in the case of those regenerated from irradiated cultures. Cytological study showed no alteration in the somatic chromosome number, which remained uniform at 2n=30+2B, in all the plants regenerated from the different subcultures and the irradiated cultures.ThesisItem Open Access Induction of Genetic Variability in Musa Sp. Var. Nendran By in Uitro Methods(Department of pomology and floriculture, College of horticulture,Vellanikkara, 1993) Mini Balachandran; P.K.ValsalakumariInvestigations were carried out at the Department of Pomology and Floriculture and Plant Tissue Culture Laboratory of the All India Co-ordinated Floriculture Improvement Project, College of Horticulture, Vellanikkara, Thrissur, during 1991-93 on the induction of variability in the banana variety Nendran (Musa AAB ‘Nendran’) by in vitro methods. Explants utilized for the study were shoot tip and eye bud for direct organogenesis through enhanced release of axillary buds and shoot tip, flower base, inflorescence axis, embryonic leaves and scalp for somatic organogenesis/embryogenesis. For culture establishment, axillary shoot initiation and in vitro rooting, different growth regulators, like NAA, 2, 4-D and 2, 4, 5-T (auxins) and BA and kinetin (cytokinins) were made use of. The plantlets produced in vitro were subjected to hardening treatments to ensure a better establishment of planted out plants and their growth parameters were studied. For shoot tip and eye bud explants, a combination of treatments involving, an initial dipping of explants in emisan (0.1 per cent) for 30 minutes followed by dipping in norfloxacin (0.1 per cent) for 30 minutes followed by dipping in norfloxacin (0.1 per cent) for 30 minutes and finally rinsing the explants in mercuric chloride (0.1 per cent) for 20 minutes was found to be best, but for flower base and inflorescence axis explants, emisan (0.1 per cent) treatment for 20 minutes and for embryonic leaves, dipping in alcohol for one second were in the best. Better and speedier establishment and growth of shoot tip and eye bud explants were observed on MS (semi-solid) medium containing NAA 2 ppm + BA 5 ppm. Addition of activated charcoal (500 mg per litre) to the medium, reduced media and explant discolouration due to polyphenol oxidation. When the performance of the shoot tip and eye bud explants was compared, eye bud explants took more time for culture establishment and growth. In shoot tip culture, on an average, each explant released 8.66 axillary shoots in the treatment involving MSb*+ NAA 2 ppm + BA 10 ppm. In the case eye bud, on an average, each explant released five axillary shoots. Continuous sub culturing was carried out at two week interval to assess the variation induced to cultured plants due to repeated subculturing. It was found that, the number of shoots produced per culture was not constant in all the subcultures. Still, the axillary shoots produced per explant per culture vessel increased at the mean rate of 5.90. BA alone at higher concentration (10 ppm) resulted in colloid (globular semi-hard, light green callus like structure) formation and subsequent regeneration. MSb*: MS medium containing half concentration of inorganic salts and full concentration of organic growth factors. For in vitro rooting, MSb medium containing NAA 10 ppm and AC 0.05 per cent was found to be effective for early root initiation and the maximum number of roots per shoot was produced at the treatment involving MSa* +NAA 5 PPM+ AC 0.05 per cent. Of the various explants, viz., shoot tip, inflorescence axis, flower base, embryonic leaves and scalp(in vitro) tried for initiating collus scalp and embryonic leaves recorded maximum response. Among the media tried for callus initiation, MSb media at liquid consistency was found to be more effective. Maximum callus index (266) was recorded for the treatment combination involving 2, 4-D 7 ppm and BA I ppm. For callus differentiation the treatments involving 2, 4-D and BA, BA alone and basal MS media resulted in rhizogenesis, and treatments involving 2, 4-D alone produced embryoid like structures from scalp callus. No shoot organogenesis was observed. Also treatments were conducted with changed levels of nitrate source in the media, but they did not give any favourable results. Embryoid like bipolar structures were recovered from scalp callus when they were transferred to media devoid of growth regulators. To study the variation, if any, induced due to derail subculturing, the shoots obtained from each subculture cycle (through enhanced release of axillary buds) were isolated and their MSa* : MS medium containing full concentration of inorganic salts and organic growth factors identify maintained. The shoots thus separated were rooted and planted out after subjecting them to a process of hardening. Somatic chromosome counts were made at the root tips of plantlets from 10 subcultures to confirm the ploidy. All the plants were triploids (2n = 33). The plantlets from different subcultures were planted out in sand, which was found to be the best medium. Observations made on growth parameters, at fifteen days interval, revealed that the plants from subcultures differed significantly with respect to the rate of growth in height and leaf area.