Browsing by Author "Sharma, Manju"
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ThesisItem Open Access Development of hybrid(s) of agaricus bisporus (Lange) Imbach and their evaluation for higher yield(YSPU, 2012) Sharma, Manju; Suman, B.C.The present investigations entitled “Development of hybrid(s) of Agaricus bisporus (Lange) Imbach and their evaluation for higher yield” were undertaken to isolates the homokaryons from different parental strains, their identification by fruiting trials and by ISSR markers, matings compatible homokaryons to produce hybrids, spawn preparation and production efficiency including other aspects like physiological studies and molecular characterization of developed hybrids. one thousand six hundred forty two single spore isolates were raised from parental strains namely A-15, S 11, S140 and U3 of Agaricus bisporus. These 1642, single spore isolates showed fluffy, appressed and strand types of growth. Out of those only 36 single spore isolates (nine from each parent strains to maintain uniform pattern) which were supposed to be homokaryons on the basis of slow mycelial growth rate and appressed types ofcolony morphology were selected for future studies. All single spore isolates with appressed types of colony morphology produced no fruit bodies in any replication. However the strandy slow growing single spore isolates produced fruit bodies in all replication. For the molecular characterization of selected monokaryons from each parents, six inter simple sequence repeats (ISSR) marker were used. In the ISSR fingerprints of putative single spore isolates from A-15, S11, S140 and U3 strains of Agaricus bisporus, homokaryons lacked amplification products at multiple loci; these grows slowly and all of them had appressed colony morphology. Primers P3 in A-15, P39 in S11, P30 in S-140 and P8 in U3 strains of Agaricus bisporus were proved to be the best due to the sharpness ofbanding patterns. Out of the two hundred fifty three (253) crosses, only seven crosses between theparents were compatible because compatible colonies got fused with each other and at the point of their contact some raisedgrowth was observed whereas in case of incompatible colonies in Petri plate, growth usually stoped as colonies approached near each other. Anastomosis or intermingling of compatible myceliums were also observed under the microscope. The yield studies showed that that hybrid Hb4, produced highest averageyield (26.94kg/100kg compost) and also exhibited better qualities as compared to other isolates. Four hybrids with higher production than parent strains were taken up for studying physiological requirement and molecular characterization in detail. Among various solid media studied malt extract agar media supported maximum average mycelial growth and in case of liquid media,glucose asparagine medium supported maximum average mycelia growth anddry weight and in case and was used as a basal medium for further physiological studies. The optimum temperature and pH for the growth of different hybrids and parent strains were found to be 25 O C and 6.5 respectively. Among the carbon and nitrogen sources, best growth of different hybrids including the parent strains was recorded on maltose and proline respectively onliquid as well as on solid media. In case of vitamins and growth regulators thiamine and gibberelic acid supported maximum growth of all the hybrids including the parent strains both on liquid and solid medium. RAPD studies shown that the percentage of polymorphism detected was 60.00 per cent. Maximum number of bands were produced by OPP-07 primer, while primer OPO-02yielded maximum number of unique bands in hybrid Hb4 as compared to other primers. ITS studies showed that all the isolates exhibited identical ITS length of approximately near about 650 bp. The newly developed best hybrids namely Hb1 and Hb4 showed 33.00 per cent and 31.00 per cent molecular variation respectively as compared to the commercially used strain (U3). The ITS region showed single nucleotide polymorphisms (SNPs) at six nucleotide position of 281, 290, 291,292, 293, 295 and 300 base pair in the ITS region of the best developed hybrids (Hb1, Hb4) and parent strains namely A-15, U3 of A. bisporusThesisItem Open Access Isolation, characterization and evaluation of antagonistic potential of Trichoderma against Rhizoctonia solani Kühn causing aerial blight of soybean(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2019-06) Singh, Priya; Sharma, ManjuThe Trichoderma species are of great economic importance due to their ability to control a wide array of plant pathogens. In the present investigation fifteen isolates of Trichoderma were isolated from the rhizospheric soils of soybean, rice and mungbean from various locations of Pantnagar and Dehradun. The isolates were characterized based on their cultural and morphological characteristics. The cultural characteristics included linear growth, colony colour, formation of concentric rings, pigmentation and time taken for sporulation. The morphological characters studied were shape, disposition of phialides, conidial shape, colour and formation of chlamydospores. The isolates were incubated at different temperature regimes ranging from 5 to 40°C to study the effect of various temperatures. There was no growth at 5°C and 40°C. However at temperatures 10, 15, 20, 30 and 35°C, the isolates PT-7, PT-7, DDNT-4, DDNT-4 and PT-10 respectively showed the maximum average linear growth rate (ALGR). While at 25°C, the maximum ALGR (29.83 mm/day) were recorded for the isolates PT-1, PT-8, PT-9, DDNT-4 and DDNT-5. Significantly maximum growth of all the isolates were recorded at 25°C followed by 20°C and 30°C. However, the isolates DDNT-4 and 5 respectively showed 54.60 mm and 54.27 mm colony diameter in three days. The isolates were tested in lab as well in glasshouse for the evaluation of their antagonistic potential against Rhizoctonia solani. They were tested in lab by dual culture, volatile assay and culture filtrate assay. It was observed that in dual culture method isolates PT4, PT-5, PT-8 and DDNT-5 showed complete mycelial parasitization of R. solani at 5 DAC. However in case of volatile compounds maximum inhibition per cent was shown by DDNT-2 (60.74%) while in case of culture filtrate the maximum inhibition per cent was exhibited by isolate PT-10 (71.11%). The glasshouse results revealed that PT-10 (63.67%) was significantly most effective over other treatments in suppressing the aerial blight of soybean. These isolates of Trichoderma were further evaluated for their growth promotion activity under lab and glasshouse conditions. The results obtained from lab studies clearly indicated that PT-2 (15.50 cm) recorded maximum plumule length while DDNT-4 (15.50 cm) showed highest radical length. However, under glasshouse condition DDNT-4 (34.50 cm) recorded maximum shoot length and PT-5 (19.00 cm) showed maximum root length. On the basis of the molecular identification PT-10 and DDNT-4 were identified as Trichoderma harzianum. Overall, PT-10 was found best isolate and can be exploitated in future for enhancing crop yield and suppressing aerial blight of soybean.ThesisItem Open Access Studies on isolation, identification, characterization and evaluation of potential rhizobacterial isolates against Rhizoctonia solani Kühn of soybean(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2020-11) Idong, Stanzin; Sharma, ManjuA total of thirty-one rhizobacterial isolates obtained from maize and soybean rhizospheric soil samples from various location of Pantnagar on two different media Kings B and Nutrient agar media were culturally identified and evaluated for their antagonistic potential against Rhizoctonia solani of soybean. The cultural characteristics included colony colour, shape, size, margin, surface and elevation. Seven isolates which inhibited growth of Rhizoctonia solani with highest being recorded with isolate MR3 (82.05%) followed by SR1 (68.28%), SRP3 (59.82%) , SRP4 (51.28%) , SRP1 (49.56%) , SR2 (48.72%) and SRP2 (40.18%) were morphologically, biochemically and functionally characterized. The morphological test clearly revealed that only two were gram positive and rest were gram negative. Biochemical test includes Catalse test, Oxidase test, Urease test, Starch hydrolysis test, Gelatin liquefaction, Cellulase test, Methyl red test and Citrate utilization test. On the basis of morphological and biochemical characterization it can be presumed that isolates SR1 and MR3 were related and other isolates SRP2, SR2, SRP3, SRP1 and SRP4 were more related. Screening for their functionality showed that 6 of the isolates produced nitrogen fixation, 3 exhibited phosphorus solubilization with SRP 4 showing highest solubilization index (2.50), 6 potassium solubilization with SRP 3 showing highest solubilization index (2.91), 3 exhibited siderophore production, 2 known for IAA production and 6 showed Ammonia production property. On the basis of functional and antagonistic evaluation four isolates were selected SR1, SRP3, SRP4 and MR3. The compatibility among the rhizobacterial isolates and with Trichoderma isolates (PT-6, PT-10, DDNT-4) were screened. Isolate SRP4 was compatible with all other isolates and isolate SRP3 was compatible with all Trichoderma isolates. Bacterization with potential isolates offered protection against aerial blight under controlled glass house condition with maximum growth inhibition by SRP 4 (55.00%). The potential isolates were further evaluated for their growth promotion activity and induction of defence related enzymes. The results obtained indicated that isolate SRP 4 bacterized plant showed highest shoot length (43.65 cm), root length (10.57 cm), fresh shoot weight (2.20 g) and fresh root weight (0.34 g) and chlorophyll content ( chl a-1.91mg/g FW, chl b-0.94 mg/g FW, total chl-2.85mg/g FW). Treatments that were bacterized and challenged with pathogen showed higher accumulation of POD (132.24 nmol/min/mg protein), SOD (10.27 unit/mg protein/min), Chitinase (140.56 μg glucose/min/mg protein) and β-1,3-glucanase (320.63 μg glucose/min/mg protein) as compared to only bacterized plants. While the absolute control depicted the lowest accumulation of PR proteins. Treatment SRP4 showed highest production of protein, Chitinase, β-1,3-Glucanase, POD and SOD. Overall isolate SRP4 is one such isolate that proved outstanding for suppressing aerial blight of soybean and having PGP traits also.