Browsing by Author "NATH, A.K."
Now showing 1 - 4 of 4
Results Per Page
Sort Options
ThesisItem Open Access IN VITRO SELECTION AND BIOCHEMICAL CHARACTERIZATION Of CARNATION( DIANTHUS CARYOPHYLLUS l.) CALLUS CULTURE TOLERANT OF ALTERNARIA DIANTHI(UHF,NAUNI, 2004) MEHTA, RUPALI; NATH, A.K.Carnation (Dianthus caryophyllus L.) is one of the most important cut flowers of the world and commands a respectable status. With the increase in its cultivation the limiting factors in the production of quality flowers are also increasing Among the various fungal pathogens attacking this crop, Alternaria dianthi is known to cause leaf blight of carnation. Therefore, under present investigations attempts were made to produce resistant carnation calli and characterized them by using biochemical parameters. For this an efficient protocol for establi shment of callus culture and organogenesis were standardized. For callus establishment equal concent ration (2 mg/I) of NAA and ki netin was used and for regeneration of shoots from ca Iii 0.2 mg/I BAP and 0.5 mg/I NAA was used. For rooting liquid MS medium supplemented with 2 mg/I IBA and 0.2 per cent activated charcoal was found to be best. For in vilru selection technique di fferent concentrations (5-25%) of culture fi ltrate of Alternaria dianthi was used. Selective dose of culture fi ltrate was found to be 15 per cent \vhere l 1.67 per cent calli survived. The selected calli were cultured on medium devoid of culture fi ltrate for 4 weeks. Stability fo r resistance was checked by reculturing it on selective medium. Selected calli showed significantly higher phenol s, reducing sugars, total sugars and proteins. Difference in banding pattern of isoenzyme viz . est erase, peroxidase a.1d polyphenol oxidase was obtained in the selected call iThesisItem Open Access STUDIES ON GENETIC STABILITY OF MICROPROPAGATED STRAWBERRY (Fragaria x ananassa Duch.) cv. SWEET CHARLIE(UHF,NAUNI, 2018-02) THAKUR, AAYUSHEE; NATH, A.K.ABSTRACT The present investigation was carried out to develop an efficient micropropagation system for runner tips of virus tested strawberry cv. Sweet Charlie. The runner tips were sterilized with HgCl2 for 2 minutes and established in MS medium supplemented with 0.5 mg/l BAP. Highest response of shoot multiplication was obtained in MS medium containing 0.50 mg/l BAP, 1.00 mg/l GA3 and 0.30 mg/l IBA. The multiplied shoots were rooted in MS basal medium supplemented with different concentrations of IBA. The highest average root length and average number of roots were produced in MS medium containing 0.50 mg/l IBA. The plantlets, thus developed were hardened and successfully established in a mixture of sand, soil and cocopeat in the ratio 1:1:1.In order to obtain true to type plantlets with the aid of tissue culture, it was quite necessary to examine the genetic uniformity of in vitro raised plants. Hence, in conclusion the two kinds of markers that is RAPD and ISSR markers can be successfully used to determine the genetic uniformity of in vitro raised plants. The protocol thus developed for micropropagation of strawberry cv. Sweet Charlie can be used over a long period of time without risk of somaclonal variations.ThesisItem Open Access STUDIES ON ISOLATION AND CHARACTERIZATION OF TRYPSIN INHIBITOR (TI) GENE FROM Dolichos biflorus L. (Kulth)(2013) REENA KUMARI; NATH, A.K.ABSTRACT Protease inhibitors are one of the most promising agents that confer resistance in plants against insect pests by inhibiting larval midgut proteases. Maximum extraction of trypsin inhibitor protein from seed flour of Dolichos biflorus L. was in 0.1 M phosphate buffer (pH 7.6) after four hours of extraction. Screening of Dolichos biflorus L. cultivars for trypsin inhibitor activity revealed maximum activity in HPK4 cultivar and further studies were conducted in this cultivar. Crude trypsin inhibitor of all cultivars inhibited midgut protease of P. brassicae larvae. Inhibitor activity was detected at early stages of seed development (3 days after flowering (DAF)) and it increased progressively with seed development (21 DAF to 60 DAF). Trypsin inhibitor activity decreased during seed germination as compared to dry seeds. Crude trypsin inhibitor extracted from developing and germinating seeds also inhibited larval midgut protease of S. littoralis. Neonate larvae of P. brassicae fed on cabbage leaf discs coated with 0.025-2.50 mg crude trypsin inhibitor caused 10–80 % larval mortality. The calculated LC50 value was 1.05 mg crude trypsin inhibitor and for 2.5 mg crude trypsin inhibitor the calculated LT50 value was 3.2 days. Leaf area eaten and faecal matter produced by treated larvae were significantly lower as compared to untreated controls. Larvae fed on leaf discs coated with 2.5 mg crude trypsin inhibitor for 5 days had significantly less total soluble protein in faecal matter and midgut trypsin activity as compared to untreated control. Significant reduction in egg hatching (75%) was observed in egg mass treated with 5.3mg of crude trypsin inhibitor of mature seeds. Trypsin inhibitor gene (309 bp) was amplified from cDNA synthesized from mature seeds of Dolichos biflorus L. HPK4 cultivar using designed primers. The amplified PCR product was cloned and sequenced. Sequence of Dolichos biflorus L. HPK4 cultivar trypsin inhibitor (DbTI) gene has been submitted to NCBI with Accession No. JQ259858. DbTI gene and its deduced amino acid sequence showed homology with Bowman-Birk inhibitors of Dolichos spp., Phaeolus spp., Vigna spp. and Glycine spp. The predicted molecular weight of deduced amino acid sequence was ~11.5 KDa and it had N terminal signal peptide of 19 amino acid residues. The secondary structure of deduced amino acid sequence of DbTI showed dominance of coils and sheets over alpha helix. Homology modelling was employed to predict the three dimensional structure of DbTI. Docking of trypsin enzyme and DbTI showed the inhibitor to be of non- competitive type.ThesisItem Open Access STUDIES ON PRODUCTION AND CHARACTERIZATION OF TANNASE (TANNIN ACYL HYDROLASE)(2015) NISHA; NATH, A.K.ABSTRACT Penicillium crustosum AN3 isolated from apple orchard soil is the first report from Penicillium genera to produce maximum tannase which is an important industrial enzyme used in biodegradation and food industry. A total of eleven fungal isolates were obtained from apple orchard soil, pine forest soil and botanical garden soil, respectively. Out of these eleven isolates five isolates viz., three isolates from apple orchard soil, one isolate from botanical garden soil and one isolate from pine forest soil were showed zone of degradation on Czapdox minimal medium supplemented with 0.5% tannic acid confirming tannase activity. These isolates were rescreened qualitatively for maximum tannase production in submerged fermentation isolate A3 exhibited maximum tannase activity of 2.48Uml-1. Molecular characterization of these isolates was carried out using 18S rrna gene technology and in silico analysis of 18S rrna gene sequences lead to identification of these fungal isolates as Fusarium redolens AN1, Aspergillus fumigatus AN2, Penicillium crustosum AN3, P.restrictum AN4 and P. commune AN5. Tannase producing bacterial isolates were also obtained on nutrient agar (NA) medium supplemented with 0.5% tannic acid. A total of four bacterial isolates from tea garden soil, pine forest soil, ruminial fluid and sheep excreta were screened on the basis of their ability to utilize tannic acid and were characterized morphologically, biochemically and for tannase activity leads to identification as Klebsiella sp, Enterobacter sp, Staphylococcus sp and E.coli respectively. Molecular characterization two bacterial isolates was carried out using 16S rrna gene technology and in silico analysis of 16S rrna gene sequences lead to confirmation of these isolates as Klebsiella variicola AN1 and Enterobacter hormaechei AN2. Penicillium crustosum AN3 produced maximum tannase (2.45Uml-1) in submerged fermentation was selected for tannase production in SSF. The enzyme was partially purified from culture extract of enzyme. Cultural conditions and process parameters i.e. type of substrate, incubation time, temperature, initial pH, moisture level, substrate concentration, tannic acid concentration, carbon sources, peptone, yeast extract, urea, NH4NO3 were optimized using one factor at a time approach and activity obtained was 39.5Ug-1. Further optimization was carried out using central composite design following response surface methodology with three independent variables which resulted in increased in tannase production to 55.55 Ug-1. The enzyme was partially purified by acetone precipitation and gel filteration chromatography which showed 46.48 yield with 3.94 fold purification and had a molecular mass of 205kDa. Gallic acid the hydrolytic product in crude enzyme was detected by TLC, FTIR and HPLC using gallic acid as standard. Crude enzyme obtained was studied for its ability in pine needle degradation, tea colour decolourization, dye decolourization and removal of apple browning