Browsing by Author "Deepa, S Nair"
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ThesisItem Open Access Cryoconservation of koovalam (Aegle marmelos L.Corr.) by encapsulation-dehydration technique(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2018) Deepa, E; KAU; Deepa, S NairThe study entitled “Cryoconservation of Koovalam (Aegle marmelos L. Corr.) by encapsulation-dehydration technique,” was carried out in the Department of Plant Biotechnology, College of Agriculture, Vellayani during 2015-2017. The objectives of the present study were to standardize a cryopreservation protocol for Aegle marmelos using encapsulation and dehydration technique and to assess the genetic fidelity of plantlets regenerated from encapsulated axillary buds after cryostorage, using molecular markers. Investigation was carried out in three phases viz., enhancement of multiplication rate, standardization of in vitro conservation using the encapsulation-dehydration technique and assessment of genetic fidelity of the regenerated plantlets using ISSR markers. Single node segments with axillary buds from in vitro maintained cultures were used as the explants in all the experiments. Among the twelve combinations of auxins and cytokinins were tried, MS (Murashige and Skoog, 1962) medium supplemented with BA 2 mg L-1 + IBA 0.5 mg L-1 was found to be the best treatment with respect to shoot multiplication (9.33 shoots per explant). The additives chitosan (CH), thidiazuron (TDZ) and adenine sulphate (AdS) at different concentrations were supplemented in two different media i.e. 1) hormone free MS medium and 2) MS + BA 2 mg L-1 + IBA 0.5 mg L-1 to study their effect on shoot proliferation. The best shoot proliferation response obtained for each additives were MS + BA 2 mg L-1 + IBA 0.5 mg L-1 + chitosan 10 mg L-1 (35.66 shoots per explant), MS + AdS 60mg L-1 (5.33 shoots per explant) and MS + BA 2 mg L-1 + IBA 0.5 mg L-1 + TDZ 0.02 mg L-1 (20.00 shoots per explant). Even though higher shoot proliferation was observed in CH and TDZ supplemented media, they exhibited morphological abnormalities. Normal shoots were obtained with AdS supplemented medium, but shoot proliferation was less compared to MS + BA 2 mg L-1 + IBA 0.5 mg L-1. Hence MS + BA 2 mg L-1 + IBA 0.5 mg L-1 was used as basal medium for cryopreservation studies. Encapsulation-dehydration technique of cryopreservation involved various steps like preconditioning, encapsulation, pre-culture, dehydration (desiccation), thawing and recovery. Nodal segments with single axillary buds were used as the explant. In preconditioning experiment, PC2 (sucrose 0.1M in semi solid MS for 7 days) was selected as the best preconditioning treatment, which produced maximum shoot proliferation (5.50 shoots per culture) when cultured on basal medium. Among different encapsulation treatments, maximum shoot proliferation of (6.66 shoots per culture) was obtained in the beads formed with sodium alginate 3.5 per cent in modified MS medium and calcium chloride 100 mM, when cultured on modified basal medium (½ MS + BA 2 mg L-1 + IBA 0.5 mg L-1). Pre-culture experiments were conducted using preconditioned and encapsulated explants. The pre-culture treatment selected was liquid MS medium supplemented with sucrose 0.5 M and DMSO 3 per cent for 3 days, which gave maximum shoot proliferation (3.66 shoots per explant) in modified basal medium. The preconditioned, encapsulated and pre-cultured beads were subjected to 0 to 7 h of desiccation under laminar airflow. The moisture content declined from 82 per cent to 12.60 percent on 7 h of desiccation. The desiccated beads were then cryopreserved in liquid nitrogen for 2h, followed by thawing for 30-60s at 40oC and inoculated on to recovery medium ½ MS + BA 2 mg L-1 + IBA 0.5 mg L-1. Survival (66.67 per cent) and regeneration (50 per cent) could be obtained only at 6h of desiccation when the moisture content was 19.50 per cent. The beads when stored in liquid nitrogen for different duration did not show any significant variation with respect to survival and regeneration. The genetic fidelity of plantlets regenerated from encapsulated axillary buds subjected to cryostorage were analysed using ISSR markers. Among the nine primers tried, four primers UBC 807 (AGAGAGAGAGAGAGAGT), UBC 840 (GAG AGA GAG AGA GAG AYT), UBC 847 (CACACACACACACACART) and UBC 826 (ACACACACACACACACC) that gave scorable (5-7) bands were selected for the study. The ISSR banding patterns of the cryoregenerated plantlets and control plants were identical, which indicated the genetic stability. This study was successful in developing a protocol for cryopreservation using encapsulation-dehydration technique in A. marmelos with 50 per cent regeneration efficiency.ThesisItem Open Access Establishment of in vitro regeneration systems from callus and protoplast in capsicum frutescens L.(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2018) Jancy, J Sathyaraj; KAU; Deepa, S NairThe present study entitled “Establishment of in vitro regeneration systems from callus and protoplast in Capsicum frutescens L. was carried out in the Department of Plant Biotechnology, College of Agriculture, Vellayani during 2015-2017. The objective of the study was to establish callus culture from different explants in C. frutescens, to establish protocol for protoplast isolation from callus/leaf mesophyll and to culture protoplast. The study was carried out in two phases viz., establishment of callus culture and organogenesis; and standardization of protocol for protoplast culture. Callus was induced from leaves and internodal segments from in vitro raised seedlings. Among the 44 treatments in MS medium with different combinations of auxins (NAA, IAA, IBA, 2,4-D and picloram) and cytokinins (BA and Kn), 100 per cent callus induction was obtained in MS media supplemented with picloram 0.50, 1.00, 1.50 and 2.00 mg L-1 (C41, C42, C43, and C44), NAA 1.50 mg L-1, NAA 2.00 mg L-1(C3, C4) and BA 3.00 mg L-1 + NAA 1.00 mg L-1 (C29). Among the 78 treatments tried for organogenesis, calli obtained from C29 treatment showed organogenesis in MS + BA 3.00 mg L-1 + IBA 1.00 mg L-1 (R37) and (MS + BA 5.00 mg L-1 + IAA 2.00 mg L-1 (R61) in 41 and 90 days, respectively. The microshoots obtained recorded 83.33 per cent rooting in MS medium supplemented with IAA 1.50 mg L-1(Rt7) in 12.83 days. Leaves excised from in vitro seedlings, and calli produced in MS + BA 3.00 mg L-1 + NAA 1.00 mg L-1 (C29), were used as explants for protoplast isolation. Leaf bits incubated in cell protoplast washing (CPW) solution containing cellulase 2.00 per cent, macerozyme 0.50 per cent and mannitol 0.50 M (maintained at pH 5.8) for 6 h (DM28) in dark at 27oC, yielded (124 x 105) protoplast per g, with a viability of 95.16 per cent. The callus yielded maximum protoplast (36 x 105 protoplasts per g) in an enzyme combination of cellulase 2.00 per cent + macerozyme 0.50 per cent + mannitol 0.60 M (maintained at pH 5.8) after 4 h 91 (DM47) of incubation under same conditions. In protoplast purification, floatation medium with 21 per cent sucrose recorded maximum protoplast yield (30 x 105 protoplasts per g tissue and 10 x 105 protoplasts per g callus) and maximum viability (90.91 per cent and 100 per cent), from leaf derived and callus derived protoplast, respectively. The purified protoplasts with 10 x105 plating density initiated microcalli in liquid MS medium supplemented with mannitol 0.50 M, 2,4- D 0.50 mg L-1 and sucrose 30g L-1 (PCM5) in 45 days. Further development to visual colony formation of microcalli was obtained on addition of liquid MS medium supplemented with mannitol 0.40 M and sucrose 5g L-1, in 60 days from callus derived protoplast and in 70 days from leaf derived protoplast. In the study, maximum callusing response was obtained in MS medium with picloram 1.50 mg L-1. Organogenesis was obtained from the calli derived in MS medium with BA 3.00 mg L-1 and NAA 1.00 mg L-1. The shoot initiated from the calli in MS medium with BA 3.00 mg L-1 and IBA 1.00 mg L-1. The rooting of microshoots could be obtained in MS medium with IAA 1.50 mg L-1. In protoplast isolation, leaf gave higher protoplast yield and viability in CPW solution with cellulase 2.00 per cent, macerozyme 0.50 per cent and mannitol 0.50 M, incubated in dark for 6 h and callus, in CPW solution with cellulase 2.00 per cent, macerozyme 0.50 per cent and mannitol 0.60 M, incubated in dark for 4 h. The protoplasts purified in 21 per cent sucrose supplemented floatation medium and adjusted to a plating density of 10 x 105, initiated microcalli in liquid MS medium supplemented with mannitol 0.50 M, 2,4- D 0.50 mg L-1 and sucrose 30g L-1. The visual colony formation of microcalli was obtained on addition of liquid MS medium supplemented with mannitol 0.40 M and sucrose 5g L-1. In this study, a callus mediated in vitro regeneration system has been established in C. frutescens. A protocol has also been developed for protoplast isolation from leaf and calli, and its culture resulting in microcalli formation.ThesisItem Open Access Evaluation of genetic stock of sanghupushpam (Clitoria ternatea L.) for yield, Alkaloid content and nitrogen fixing potential(Department of Horticulture, College of Agriculture, Vellayani, 2000) Deepa, S Nair; KAU; Reghunath, B RThe present study entitled “Evaluation of genetic stock of ‘Sanghupushpam’ (Clitoria ternatea L.) for yield, alkaloid content and nitrogen fixing potential” was carried out at the Department of Horticulture, College of Agriculture, Vellayani from June 1999 to January 2000. Seeds of twenty different accessions were collected from various locations from inside and outside the state. Thirteen accessions having high rate of seed germination were raised as intercrop in young coconut garden and maintained till seed maturation stage. The performance of the accessions in terms of growth, yield and physiological parameters were evaluated. Growth and yield parameters with respect to shoot, pod, root and root nodule characters were evaluated. Physiological parameters evaluated included leaf area index, leaf area duration, net assimilation rate, crop growth rate, relative growth rate, absolute growth rate and harvest index. Number of effective nodules was taken as an index for assessing nitrogen fixing potential of the plant. Leaf yield, shoot yield and root yield were significantly superior in the accession MP-90. MP-83 recorded significantly superior pod yield. The number of nodules was the highest in accessions MP-76 and MP-82. Crude alkaloid content was significantly superior in seeds of MP-74 and MP-76. Six accessions were selected based on yield , nodule characters and crude alkaloid content viz ., MP-90, MP-74, MP-83, MP-78, MP-76 and MP-82. Results of the present study indicated that Clitoria ternatea, is not a good proposition as an intercrop in young coconut garden. However, it may be worth studying the performance of the crop as a pure crop under open condition or as an intercrop in coconut gcirden with comparatively lesser shade (less than 50 per cent) than the situation of the present study.ThesisItem Open Access Growth, yield and essential oil production responses to microbial elicitation in Ocimum basilicum L.(Department of Plantation Crops and Spices, College of Agriculture, Vellayani, 2021) Rajeswari, E; KAU; Deepa, S NairThe seeds of O. basilicum used for the study were sourced from Anand Agricultural University, Gujarat. The study was carried out in two phases: Phase 1- Seed priming using fungal derivatives for enhanced germination. Phase 2- Evaluation of the effect of foliar application of fungal derivatives for growth, yield and essential oil production. In the first phase of study, the seeds were subjected to various priming treatments using fungal derivatives viz., Trichoderma viride cell wall extract (1 %) (TCWE), Trichoderma viride culture filtrate (1 %) (TCF), Piriformospora indica cell wall extract (1 %) (PCWE), Piriformospora indica culture filtrate (1 %) (PCF) and hydro priming, maintained upto 30 days after sowing. The seeds without any priming were taken as the absolute control. In the second phase of study, the 30 days old seedlings of O. basilicum were transplanted to grow bags. The foliar spray of corresponding fungal derivatives (cell wall extract and culture filtrate) at 1 % concentration were given to plants at fortnightly intervals from transplanting to 90 days after sowing. The treatment without any foliar application was taken as the absolute control. The seeds bioprimed with PCF @ 1 per cent recorded the highest germination per cent (96%), survival per cent (96%) and had taken minimum number of days (3 days) to initial sprouting. While TCF @ 1 per cent exhibited the highest germination index (34.50) and lowest mean germination time (6.29 days). With regard to seedling development, PCF @ 1 per cent recorded a significantly higher shoot length (21.50 cm), root length (19.50 cm), seedling length (41.00 cm) and seedling vigour index (39.37). The highest (1.07) allometric index was observed in the treatment PCWE @ 1 per cent. At 110 DAS, the plants subjected to foliar application with PCF @ 1 per cent exhibited higher plant height (80.20 cm), collar girth (6.03 cm), leaf area (4010.82 cm2 ), number of branches (28.00) and number of flowering shoots (104.00). The same treatment induced early flowering (55 days) in O. basilicum. The foliar spray treatment with PCF @ 1 per cent exhibited significantly higher total chlorophyll content (1.20 mg g-1 ) and polyphenol content (84.31 mg PE g-1 ) at 110 DAS. The plants subjected to foliar application with PCF @ 1 per cent recorded maximum leaf biomass (210.00 g and 19.04 g), stem biomass (135.33 g and 12.21 g), herbage yield (345.33 g and 31.25 g), root biomass (52.00 g and 4.63 g) and total plant biomass (397.33 g and 35.88 g) respectively, on both fresh weight and dry weight basis. The same treatment recorded the highest leaf biomass (125.33 g and 12.44 g), stem biomass (76.00 g and 7.31 g), and herbage yield (201.33 g and 19.75 g), on fresh weight and dry weight basis respectively, in the ratoon crop harvested 60 days after the first cut. PCF @ 1 per cent was also observed to give the highest essential oil content (2.11 per cent and 1.00 per cent) and oil yield (443.10 g and 19.04 g, respectively) in terms of both fresh and dry leaf weight. This is followed by PCWE @ 1 per cent and TCF @ 1 per cent in terms of oil content and yield. In the first phase of study, PCF @ 1 per cent gave better performance in terms of seed germination, seedling growth and seedling vigour index. The transplanted seedlings from the same treatment when subjected to foliar application with PCF @ 1 per cent at fortnightly intervals gave the highest plant growth, biochemical and yield parameters in the second phase of study. Hence, it can be inferred that biopriming followed by foliar application of the fungal derivative PCF @ 1 per cent would give superior performance in terms of plant growth, yield and essential oil production in O. basilicum.ThesisItem Open Access Growth, yield and secondary metabolite production responses to microbial elicitation in Withania somnifera (L.) Dunal(Department of Plantation Crops and Spices, College of Agriculture, Vellayani, 2021) Ragin, Shaji; KAU; Deepa, S NairThe study entitled “Growth, yield and secondary metabolite production responses to microbial elicitation in Withania somnifera (L.) Dunal.” was conducted at the Department of Plantation Crops and Spices, College of Agriculture, Vellayani, Thiruvananthapuram during 2019-2021 with a view to evaluate the effect of bacterial inoculants on seed germination, seedling vigour, growth, yield and secondary metabolite production in W. somnifera. Seeds of W. somnifera were primed with B. amyloliquefaciens (Bam), B. pumilus (Bp) and B. velezensis (Bv) at 1x 108 cfu mL-1 individually and in combination for 24 h. Among these treatments, T7, the trio combination of Bam+Bp+Bv recorded the earliest germination (5.33 days) highest germination per cent (96.67), survival per cent (92.67) seedling vigour index (958.93), basal shoot girth (0.81 cm), number of leaves (6.07), leaf area (13.38 cm2 ), shoot length (5.77cm), root length (4.16 cm) and root volume (0.54 cm3 ). All the biopriming treatments with Bacillus spp. recorded superior germination and seedling parameters over the untreated control (T9) and hydropriming (T8). The seedlings from the first phase were subjected to root dip with the respective bacterial suspension for 30 min on transplanting. The morphological and yield determining parameters such as shoot length(78.99 cm), root length (21.27cm), number of branches (8.78), number of leaves (71.00), collar girth (3.91 cm), leaf area (5146.81 cm2 ) number of flowering branches (7.89), stem fresh weight (61 .85 g plant -1 ), stem dry weight(9.78 g plant -1 ), leaf fresh weight (45.89 g plant -1 ), leaf dry weight (5.07g plant - 1 ), root fresh weight (5.47g plant-1 ), root dry weight (1.44 g plant-1 ) 100 seed weight (0.26g ) root diameter (1.33cm), root volume (5.39 cm3 ) and harvest index (0.10) were observed to be significantly higher in T7, the trio combination of (Bam+ Bp+ Bv), which was observed to be on par with T5, dual combination of (Bam+ Bv). T5 was found to be superior in shoot fresh and dry weight, berry fresh and dry weight, number of berries and 133 seed yield per plant and total dry matter production (97.48, 17.51, 8.85 and 5.32 g plant-1 , 90.56, 7.35, 18.89 g plant-1 respectively, which was observed to be on par with T7. All the said parameters were significantly lower in untreated control. Seedlings treated with bacterial suspension of B. velezensis (Bv) recorded highest chlorophyll content in the leaves of W. somnifera at the time of harvest. The highest total alkaloid content in leaves (7.86 µg 100 mg-1 ) was recorded in dual combination of Bp+Bv which was on par with the other combinantions, Bam+Bv (T5) and Bam+Bp+Bv (T7). T5 recorded the highest protein and carbohydrate content (2.96 and 23.30 mg 100 mg-1 respectively) in the roots which was on par with T7. The withanolide content was superior (7.46 µg mg-1 ) in T7, Bam+Bp+Bv which was on par with T5, Bam+Bv and T6, Bp+Bv. The yield of biochemical parameters on per plant basis viz., total leaf alkaloids, total root withanolides were the highest (397.44 µg plant-1, and 10.77 mg plant-1 respectively) in trio combination of T7 which was on par with dual combination T5. The control treatment recorded significantly lower values in all the biochemical parameters observed. In the first phase of the study, the trio combination of Bam + Bp+ Bv (T7) gave the best performance in terms of seed germination, seedling growth and seedling vigor index, In the second phase, Bam + Bp+ Bv (T7) and Bam+ Bv (T5) gave superior performance, in terms of plant growth, yield and biochemical parameters.ThesisItem Open Access In vitro conservation of chethikoduveli (Plumbago rosea L.) using encapsulation and vitrification techniques(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Sowmya, A S; KAU; Deepa, S NairThe present study entitled “In vitro conservation of chethikoduveli (Plumbago rosea L.) using encapsulation and vitrification techniques” was carried out in the Department of Plant Biotechnology, College of Agriculture, Vellayani. The objective of the study was to standardise the protocol for short term and long term conservation of axillary buds using encapsulation and vitrification techniques and to assess the genetic fidelity of recovered and regenerated plantlets using molecular markers. The study was carried out in three phases viz., enhancement of multiplication rate, short term conservation by encapsulation technique and long term conservation using vitrification technique. Axillary buds from in vitro raised cultures were used as the explants in all the experiments. Different media (Gamborg, White, SH and MS) and different levels of additives (chitosan, adenine sulphate and thidiazuron) were tried for the enhancement of multiplication. Standardized medium, MS supplemented with BA 1.5mg L-1 and IAA 1mg L-1, for shoot proliferation was used as the control treatment in the study.Among the different media tried, MS medium was observed to give the best response with respect to shoot multiplication (3.75 shoots/explant). Among the different levels of each additive tested, best response was obtained with chitosan 15 mg L-1, adenine sulphate 40 mg L-1 and thidiazuron 4 mg L-1 and 5mg L-1 in MS supplemented with BA 1.5 mg L-1 and IAA 1 mg L-1. Thidiazuron at both levels gave maximum shoot proliferation (11.5 shoots/explant). But shoots obtained were stunted, and vitrified with callusing at the base. Adenine sulphate 40mg L-1, gave significantly better response with 4.25 shoots/explant. Though chitosan 15mg L-1gave maximum shoots/explants (3.67) among different levels of chitosan tried, it was found to be on par with the control. Hence, MS medium supplemented with BA 1.5mg L-1, IAA 1mg L-1 and adenine sulphate 40 mg L-1 was selected as the medium for further conservation studies. In short term conservation studies, encapsulated axillary buds were used as explants. Effect of different additives (sucrose 10 per cent, sucrose 15 per cent, mannitol 10 per cent, mannitol 15 per cent) in encapsulation matrix (sodium alginate 2.5 per cent and calcium chloride 100 mM), different storage media (liquid MS, sterile distilled water) on shoot regeneration and proliferation were tried at two temperature regimes of 4˚C and 25˚C. Encapsulated axillary buds could be stored upto 30 days in all combinations, beyond which the regeneration percentage deteriorated. Maximum regeneration of 83.33 and 79.17 per cent was obtained with mannitol 10 per cent in encapsulation matrix and liquid MS as storage medium at 25˚C and 4˚C respectively after 30 days of storage. In this treatment at 25oC, the days to bud initiation was the least (11.60 days); Shoots/explant (6.4), shoot length (1.87cm) and nodes per shoot (2.48) was the highest. Two cryopreservation techniques of simple vitrification and encapsulation vitrification were attempted for long term conservation studies. In simple vitrification, preconditioned (0.5 M sucrose for 7 days), precultured (0.5 M sucrose for 3days) axillary buds were exposed to vitrification solutions, PVS2 (glycerol 30 per cent, ethylene glycol 15 per cent and DMSO 15 per cent in MS with sucrose 0.4 M, pH 5.7) and PVS3 (glycerol 50 per cent and sucrose 50 per cent in MS, pH 5.7) for 0 to 210 minutes at 30 minutes interval. The best response in terms of survival (62.22 per cent) and regeneration (47.78 per cent) was obtained in PVS2 exposure for 30 min after 2 h of cryopreservation ie., storage in liquid nitrogen. In encapsulation vitrification, preconditioned, encapsulated, precultured axillary buds were given exposure to vitrification solutions as above, maximum survival (78.89 per cent) and regeneration (70.00 per cent) was also obtained in the same treatment. The survival and regeneration percentage was found to be on par with different periods of cryopreservation. Among the cryopreservation treatments, encapsulation vitrification was found to be the best. The cryoregenerated explants were then inoculated on to the best proliferation medium to study the regeneration and multiplication. Encapsulation vitrification in PVS2 for 30 minutes gave best response with respect to bud initiation (28.89 days), shoots/ explant (2.56) and shoot length (2.88 cm). The genetic fidelity of the plantlets subjected to short and long term conservation was assessed using 5 RAPD and 4 ISSR markers. Among these, two ISSR primers produced 5-7 bands. The banding patterns of the conservation regenerated plantlets and the control were compared. The profiles generated did not show any polymorphism and was identical to those of control, which indicated the genetic stability. In the study, maximum multiplication rate (4.25 shoots/explant) was obtained in MS media supplemented with BA 1.5mg L-1, IAA 1mg L-1 and adenine sulphate 40mg L-1. With respect to short term conservation, the encapsulated axillary buds with the additive in encapsulation matrix (mannitol 10 per cent), storage media (liquid MS) and storage temperature (25˚C) gave maximum regeneration (83.33 per cent). Encapsulated axillary buds could be stored upto 30 days in all combinations, beyond which the regeneration percentage deteriorated. In long term conservation the maximum survival (78.89 per cent) and regeneration (70.00 per cent) was obtained in encapsulation vitrification. The genetic stability was maintained in plantlets regenerated from short and long term conservation.ThesisItem Open Access Standardisation of nursery management practices in pachotti (symplocos cochinchinensis (lour.) s. moore)(Department of Plantation Crops and Spices, College of Agriculture, Vellayani, 2018) Ajil, M S; KAU; Deepa, S NairThe study entitled “Standardisation of nursery management practices in pachotti (Symplocos cochinchinennsis (Lour.) S. Moore)” was carried out in the Department of Plantation Crops and spices, College of Agriculture, Vellayani durng 2017-18. The objective of the study was to evaluate the propagation efficiency of different propagules viz., seeds, stem cuttings and root cuttings and to standardise the potting media for the nursery plants of pachotti. The propagules viz., seeds, stem cuttings and root cuttings for the study were sourced form Jawaharlal Nehru tropical Botanical Gardens and Research Institute, Palode, Thiruvananthapuram and from Wayanad district. The seeds were subjected to in vivo and in vitro germination studies. In in vivo study, among the pretreatments tried, viz., physical treatments, chemical priming and bio priming, only physical treatment of scarification (with sand paper) responded with a very low germination of 2 per cent. The germination commenced after two months of the treatment. Other in vivo pretreatments as well as in vitro treatments did not give any germination. In vegetative propagation, stem cuttings were exposed dto hormone/chemicals (auxins, phloroglucinol and salicylic acid (SA) pretreatments for two hours before planting. When pretreated with SA @ 10 and 20 mg L -1, at three months after planting, the hardwood cuttings responded with 30 pere cent survival, whith a shoot length of 2.99 mcm and 3.62cm, respectively. The semihardwood cuttings pretreated with SA@ 20 mg L-1 responded with 23.33 per cent survival with a higher shoot length of 3.72 cm. Both the hardwood and semi hardwood cuttings pretreated with SA 20 mgL-1 had on par values with respect to shoot length. Root cuttings were pretreated with different concentrations of various types of auxins. Root cuttings pretreated with IAA @ 250 mg L -1, after three months of planting responded with 33.33 per cent survival with a shoot length of 5.73 cm. Though root cuttings had slightly higher survival percent and shoot length, hardwood cuttings were selected for the valuation of potting media due to better availability and ease in procurement. The three month old hardwood cuttings pretreated with SA @ mgL-1 were then transplanted to ten different potting media comprising of two basal media viz., soil:coipith compost :cowdung (1:1:1) (B1) and soil : soirpith compost : vermicompost (1:1:1) (B2), and each in combination with biofertilisers @ 5g plant -1 viz., PGPR (Plant Growth Promotng Rhizobacteria) Mix I, Azospirillum, PSB (Phosphorus Solubilising Bacteria ) and AMF (Arbuscular Mycorrhizal Fungi). At fourth month after transplanting, B2 in combination with biofertilisers were found to be significantly superior to B2, B1 and B1 in combination with biofertilisers with respect to morphological parameters. B2 +PGPR Mix I recorded highest shoot length (11.50 cm) and number of leaves (10.50) which was on par with B2+Azospirillum, B2+PSB and B2+AMF; the highest number of branches (1.92) was observed in B2+ Azospiriillum which was on par with the treatments , B2+ PGPR Mix I, B2+PSB and B2+AMF. The fresh and dry weight of shoots were the highest (21.35 g and 4.78 g respectively) in B2 +PGPR Mix I which was on par with B2+ AMF. B2+ AMF recorded highest values (4.77 cm, 0.30 mm, 3.28 g and 0.0092 g, respectively) with respect to root growth parameters viz., root length, root girth , fresh and dry weight of roots. The physiological parameters, leaf area index (1.36) and leaf area duration (34.63 days) were the highest in B2+ PGPR Mix I which was on par with B2 in combination with other biofertilisers. The phytochemical analysis indicated that carbohydrate content (80.9 mg g-1) of plant tissue was the highest in B2+PGPR Mix 1, which was on par with B2+PSB, B2+Azospirrillum and B2+AMF. Chlorophyll content was found to the highest (1.20 mg g-1) in B2+ Azospirillum which was on par with B2+PGPR Mix I. The same treatment recorded the highest soluble protein content (20.31mg g-1) and it was on par with B2 in combination other biofertilisers. The nutrient analysis of plant tissue showed that nitrogen (2.22 percent ) and potassium (2.15 per cent ) content was significantly higher in B2+ Azospirillum. B2+PSB Recorded higher phosphorus content (0.26 per cent) among the treatments. The study indicated that nursery plants in the potting media B2 in combination with biofertilizers gave better performance with respect to morphological parmeters, physiological parameters, phytochemicals and plant nutrients. Among the various potting media tried, B2+AMF recorded significantly higher plant growth potential (0.522) followed by B2 + PGPR Mix I (0.428). In the study, hardwood cuttings were identified as the preferred planting material for the nursery establishment of pachotti. The cuttings could be treated with salicylic acid @ 20 mg L-1 for initial establishment of nursery plants. The preferred potting media for transplanting the established cuttings for raising the nursery plants of pachotti is Soil : Coirpith compost : Vermicompost (1:1:1) +AMF (5g/plant).ThesisItem Open Access Utilization of pineapple (Ananas comosus(L) Merr.) biomass for biofuel production(College of Agriculture, Vellayani, 2014) Anoop, P; KAU; Deepa, S NairA study on utilization of pineapple (Ananas comosus (L.) Merr.) biomass for biofuel production was conducted at College of Agriculture, Vellayani, Thiruvananthapuram during the period of 2013-14. Rising concern over depleting fossil fuel and greenhouse gas resulted in a high level of interest in nonconventional fuel originating from biorenewable sources including sugars, starches and lignocellulosic materials. Lignocellulosic materials constitute a substantial renewable substrate for bioethanol production that do not compete with food production and animal feed. Pineapple waste is a promising feed stock for alcohol production due to its abundance and ease of availability. Also it is a cheap substrate for biofuel production due to low lignin content and can undergo hydrolysis steps more easily. The feed stocks were prepared by drying and grinding of pineapple peel, pineapple fruit waste and pineapple plant residue separately. This is a method of physical pretreatment used for degradation of lignocelluloses and for reduction of cellulose crystallinity. The study on moisture content of the feedstocks using gravimetric method showed that pineapple plant residue has higher moisture content followed by pineapple fruit waste and pineapple peel waste. The estimation of sugar content of different feed stocks revealed that, pineapple fruit waste have highest values of glucose, fructose, xylose and sucrose compared to the other feed stocks and this higher levels of sugar content resulted in higher ethanol production during fermentation. Total dissolved solids was found to be maximum in pineapple fruit waste. Similarly total carbohydrate was recorded maximum in pineapple fruit waste followed by pineapple peel waste and lowest value was observed in pineapple plant residue. Estimation of cellulose, hemicelluloses and lignin content of the feed stocks revealed that pineapple plant residue have maximum cellulose content followed by pineapple fruit waste and pineapple peel waste. Whereas pineapple peel waste recorded maximum hemicellulose content. Lignin content was found maximum in pineapple fruit waste. To obtain a highly efficient conversion, pre treatment was performed for three feed stocks with acid and alkali which reduce the lignin content and make the sugar molecules accessible for fermentation. Acid and alkali pretreatment of the pineapple feed stocks resulted an increase in total reducing sugar and total non reducing sugar concentrations. The increase in sugar concentration in pretreated feedstocks is due to the hydrolysis of cellulose and hemicellulose in to sugars. The acid and alkali pretreatment decreased the lignin content, but a higher percentage removal of lignin was observed with alkaline pretreated pineapple feed stocks. The biochemical characterisation of the feed stocks revealed the sugar content and fermentation potential. To find out the effect of pretreatment fermentation was carried out in untreated and pretreated feed stocks with Saccharomyces cerevisiae and Zymomonas mobilis. Fermentation of untreated feedstocks gave higher alcohol percent than pre-treated feed stocks inspite of the fact that pretreatments resulted in an increase in total reducing and non reducing sugars and a decrease in the lignin content . This may be due to the production of various inhibitors or due to high salt formation during pH adjustments of the pretreated feedstocks. The results of percent conversion rate of reducing sugar to alcohol indicated that pineapple fruit waste have higher conversion rate than other feed stocks where as the percent conversion of non reducing sugar is found to be maximum with pineapple peel waste. pH of the fermenting medium also tend to become acidic. Characterisation of feedstocks and alcohol yield after fermentation showed that pineapple fruit waste is the most amenable feedstock for alcohol production than other two. The alcohol yield (8.34 per cent) obtained with untreated fruit waste using S.cerevisiae was found to be significantly higher than all other combinations tried. For the enhancement of fermentation and subsequent alcohol yield, cellulolytic microorganism was isolated from degraded pineapple waste. It was identified as Bacillus sp. by biochemical and molecular characterisation. Three modes of enhancement of fermentation were performed with pineapple fruit waste; Single batch bioconversion, simultaneous saccharification and fermentation (SSF) and separate hydrolysis and fermentation (SHF) using Saccharomyces cerevisiae and isolated native microorganism. Single batch bioconversion was found to be the best enhancement method yielding 11.09 per cent alcohol. The decreased level of ethanol in other enhancement methods may be due to the negative interaction of Bacillus sp. with Saccharomyces cerevisiae. The present study concluded that fruit waste is the best candidate for bioethanol production than other pineapple feed stocks tried. Single batch bioconversion using the cellulolytic organism, Bacillus sp. and fermenting organism, S. cerevisiae could bring about a substantial enhancement in alcohol yield.