ABSTRACT: The inherent regeneration ability among selected varieties of pigeonpea using decapitat... more ABSTRACT: The inherent regeneration ability among selected varieties of pigeonpea using decapitated embryonal axis and stem-node explants in the presence of different growth regulators were assessed for multiple shoot bud induction. Among three different hormones namely BAP, Kinetin and TDZ tested for in vitro regeneration at different concentration for decapitated embryonal axis explants, BAP was found to be comparatively better as evident from number of buds per explants. IPA-242 variety was found to be the best for direct organogenesis resulting in the formation of 10 buds when subjected to MS Medium supplemented with 2.0 mgL-1 of BAP. Under treatment with different concentration of TDZ, Pusa-9 revealed 10 shoot buds with 0.15 mgL-1 of TDZ. The overall response of these varieties at different concentration of kinetin was very poor. With stem-node explants under variable concentration of BAP, IPA-3088 performed best showing 17 buds per explants. Similarly Pusa-9 and IPA 3088 revea...
The Dof (DNA binding with one finger) transcription factor family represents a typical example of... more The Dof (DNA binding with one finger) transcription factor family represents a typical example of zinc finger transcription factors associated with diverse functions exclusively in plants. In the present study three dimensional structure of twenty one Dof proteins of sorghum were predicted by I-TASSER server based multiple threading method and the resulting modelled structures were validated by PROCHECK, Ramachandran Plot and QMEAN Z-score. These models were assigned PMDB IDs PM0079009 to PM0079021 and PM0079023 to PM0079030. The secondary structure revealed diversity in terms of sheets, beta-alpha-beta units, beta hairpin, beta bulges, strands, helices, helix-helix interactions, beta turns and gamma turns. The structural motif scan revealed presence of specific amino acid rich profile for each SbDof proteins along with a typical zinc finger Dof type profile. The putative functions were also predicted based on gene ontology (GO) terms in I-TASSER server. In-silico expression profili...
A total of 121 protein sequences of pectate lyases were subjected to homology search, multiple se... more A total of 121 protein sequences of pectate lyases were subjected to homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. The phylogenetic tree constructed revealed different clusters based on different source organisms representing bacterial, fungal, plant, and nematode pectate lyases. The multiple accessions of bacterial, fungal, nematode, and plant pectate lyase protein sequences were placed closely revealing a sequence level similarity. The multiple sequence alignment of these pectate lyase protein sequences from different source organisms showed conserved regions at different stretches with maximum homology from amino acid residues 439-467, 715-816, and 829-910 which could be used for designing degenerate primers or probes specific for pectate lyases. The motif analysis revealed a conserved Pec_Lyase_C domain uniformly observed in all pectate lyases irrespective of variable sources suggesting its possible role in structural and enzym...
An acidic polygalacturonase (PG) secreted by Rhizopus oryzae MTCC-1987 in submerged fermentation ... more An acidic polygalacturonase (PG) secreted by Rhizopus oryzae MTCC-1987 in submerged fermentation condition has been purified to electrophoretic homogeneity using ammonium sulphate fractionation and anion exchange chromatography on diethylaminoethyl cellulose. The purified enzyme gave a single protein band in sodium dodecyl sulphatepolyacrylamide gel electrophoresis analysis with a molecular mass corresponding to 75.5 kDa. The K m and k cat values of the PG were 2.7 mg/mL and 2.23 × 103 s−1, respectively, using citrus polygalacturonic acid as the substrate. The optimum pH of the purified PG was 5.0 and it does not loose activity appreciably if left for 24 hours in the pH range from 5.0 to 12.0. The optimum temperature of purified enzyme was 50°C and the enzyme does not loose activity below 30°C if exposed for two hours. The purified enzyme showed complete inhibition with 1 mM Ag+, Hg2+ and KMnO4, while it was stimulated to some extent by Co2+. The purified PG exhibited retting of Cro...
Genetic Engineering and Biotechnology Journal, 2010
The genetic relatedness of three varieties of finger millet (Eleusine coracana L.) with varying s... more The genetic relatedness of three varieties of finger millet (Eleusine coracana L.) with varying seed coat color namely PRM-1 (Brown), PRM-701 (White) and PRM-801 (Golden) were assessed with 10 RAPD and 10 ISSR markers. Molecular parameters viz., total number of bands, average polymorphic bands, average percent polymorphism, average polymorphic information content (PIC) and average expected gene diversity (Hi) generated from RAPD and ISSR markers has been compared. The RAPD profiling of ...
Physiology and molecular biology of plants : an international journal of functional plant biology, 2016
Nuclear factor Y (NF-Y) is a heterotrimeric transcription factor (TF) complex with preferential b... more Nuclear factor Y (NF-Y) is a heterotrimeric transcription factor (TF) complex with preferential binding to CCAAT elements of promoters, regulating gene expression in most of the higher eukaryotes. The availability of plant genome sequences have revealed multiple number of genes coding for the three subunits, namely NF-YA, NF-YB and NF-YC in contrast to single NF-Y gene for each subunit reported in yeast and animals. A total of 33 NF-YTF comprising of 8 NF-YA, 11 NF-YB and 14 NF-YC subunits were accessed from the sorghum genome. The bioinformatic characterization of NF-Y gene family of sorghum for gene structure, chromosome location, protein motif, phylogeny, gene duplication and in-silico expression under abiotic stresses have been attempted in the present study. The identified SbNF-Y genes are distributed on all the 10 chromosomes of sorghum with variability in the frequency and 18 out of 33 SbNF-Ys were found to be intronless. Segmental duplication event was found to be predominan...
An extracellular pectin lyase secreted by Fusarium decemcellulare MTCC 2079 under solid state fer... more An extracellular pectin lyase secreted by Fusarium decemcellulare MTCC 2079 under solid state fermentation condition has been purified to electrophoretic homogeniety by using ammonium sulfate fractionation, carboxymethyl cellulose and gel filtration (Sephadex G-100) column chromatographies. The purified enzyme showed single protein band corresponding to molecular mass 45 ± 01 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme had maximum activity at pH 9.0 and showed maximum stability in the pH range of 9.0–12.0. The optimum temperature of the purified enzyme was 50 °C and it showed maximum stability upto 40 °C. The energy of activation for the thermal denaturation (Ea) was 59.06 kJ mol−1 K−1. The Km and kcat values using citrus pectin as the substrate were 0.125 mg ml−1 and 72.9 s−1 in 100 mM sodium carbonate buffer pH 9.0 at 50 °C. The biophysical studies on pectin lyase showed that its secondary structure belongs to α + β class of protein with comparativ...
Heterotrimeric nuclear factors Y (NF-Ys) are involved in regulation of various vital functions in... more Heterotrimeric nuclear factors Y (NF-Ys) are involved in regulation of various vital functions in all eukaryotic organisms. Although a number of NF-Y subunits have been characterized in model plants, only a few have been functionally evaluated in crops. In this work, a number of genes encoding NF-YB and NF-YC subunits were isolated from drought-tolerant wheat (Triticum aestivum L. cv. RAC875), and the impact of the overexpression of TaNF-YB4 in the Australian wheat cultivar Gladius was investigated. TaNF-YB4 was isolated as a result of two consecutive yeast two-hybrid (Y2H) screens, where ZmNF-YB2a was used as a starting bait. A new NF-YC subunit, designated TaNF-YC15, was isolated in the first Y2H screen and used as bait in a second screen, which identified two wheat NF-YB subunits, TaNF-YB2 and TaNF-YB4. Three-dimensional modelling of a TaNF-YB2/TaNF-YC15 dimer revealed structural determinants that may underlie interaction selectivity. The TaNF-YB4 gene was placed under the contro...
Bacterial, fungal, plant and animal lipases amino acid sequences retrieved from different databas... more Bacterial, fungal, plant and animal lipases amino acid sequences retrieved from different databases were analyzed for homology search, multiple sequence alignment, phylogenetic and motif assessment using different bioinformatics tools. The multiple sequence alignment revealed different conserved stretches of amino acids exclusively for each groups along with a highly conserved glycine residue for plant and animal lipases. The phylogenetic tree constructed for different groups uniformly showed two major clusters divided into several sub-clusters based on the closeness of the lipases protein sequences irrespective of the source organisms. The 64 bacterial lipases protein sequences were distributed into two main clusters with various sub-clusters revealing the rich diversity of lipases among diverse bacterial strains. The fungal lipase protein sequences based phylogenetic tree revealed two major clusters for representing two important groups namely ascomycota and basidomycota with seve...
Microbial Diversity and Biotechnology in Food Security, 2014
The genomic sequences of Aspergillus flavus NRRL 3357 revealed seven pectin lyase (pnl) genes, wh... more The genomic sequences of Aspergillus flavus NRRL 3357 revealed seven pectin lyase (pnl) genes, which were used to design gene-specific primers and these genes were amplified through polymerase chain reaction (PCR) from five strains of A. flavus, namely A. flavus MTCC 7589, A. flavus MTCC 10938, A. flavus MTCC 8836, A. flavus NIICC 8142 and A. flavus NIICC 8147. All the seven predicted pnl genes were amplified from the genomic DNA of A. flavus NIICC 8142, while six pnl genes were from A. flavus MTCC8837 and five pnl genes from the remaining strains, respectively. A total of five pnl gene sequences of A. flavus NIICC 8142 designated as Afpnl-1, Afpnl-2, Afpnl-3, Afpnl-4, and Afpnl-5 were submitted to GenBank and assigned accession numbers JQ735890 to JQ735894, respectively. These sequences were subjected to in silico characterization for homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. The homology search revealed their identity to the predicted pectin lyase genes of A. flavus NRRL 3357, and multiple sequence alignment of these five genes showed various conserved residues. The phylogenetic tree revealed two distinct clusters and four subclusters with five pectin lyase genes of A. flavus occupying distinct position among the PCR-amplified pectin lyase genes from different fungi. The presence of unique pec_lyase C domain was observed among these sequences.
A neutral pectin lyase produced by a new fungal strain Oidiodendron echinulatum MTCC 1356 under s... more A neutral pectin lyase produced by a new fungal strain Oidiodendron echinulatum MTCC 1356 under solid state fermentation using wheat bran as agro waste has been studied. The enzyme was purified by ammonium sulphate precipitation (30-60%), DEAE anion exchange and Sephadex G-100 column chromatographies. The SDS-PAGE and native PAGE revealed two bands of sizes 42 and 47 kDa. The enzyme was purified 37 fold with specific activity of 4.5 U/mg and 2.25% yield. The K(m) and V(max) values determined using citrus pectin were 1.2 mg/ml and 0.36 IU/min respectively. The pH and temperature optima were pH 7.0 and 50 °C, respectively. The pH stability was around 5.0 for 24 h at 20 °C. The purified enzyme retained maximum activity for 30 min upto 50 °C. The activation energy for thermal denaturation of the purified enzyme was found to be 60.0 kJ/Mol. The effects of various metal ions and protein inhibitors on enzyme activity have revealed total inhibition of the enzyme activity in the presence of Ag(+) and Cu(+) and KMnO(4) at 1 mM. The neutral pectin lyase showed retting of Crotalaria juncea fibre in the presence of EDTA.
The diversity of Aspergillus from soil of Kusmi Forest, an Indian subtropical Teak forest at Gora... more The diversity of Aspergillus from soil of Kusmi Forest, an Indian subtropical Teak forest at Gorakhpur resulted in characterizing a total of 415 isolates which were then subjected to amylases production. The preliminary amylases production revealed a total of 14 promising isolates of Aspergillus comprising of ten different strains of A. flavus, two strains of A. tamarii, one strain each of A. niger and A. awamori which are characterized further. The optimum temperature of amylases produced from 14 isolates ranged from 30 ...
ABSTRACT: The inherent regeneration ability among selected varieties of pigeonpea using decapitat... more ABSTRACT: The inherent regeneration ability among selected varieties of pigeonpea using decapitated embryonal axis and stem-node explants in the presence of different growth regulators were assessed for multiple shoot bud induction. Among three different hormones namely BAP, Kinetin and TDZ tested for in vitro regeneration at different concentration for decapitated embryonal axis explants, BAP was found to be comparatively better as evident from number of buds per explants. IPA-242 variety was found to be the best for direct organogenesis resulting in the formation of 10 buds when subjected to MS Medium supplemented with 2.0 mgL-1 of BAP. Under treatment with different concentration of TDZ, Pusa-9 revealed 10 shoot buds with 0.15 mgL-1 of TDZ. The overall response of these varieties at different concentration of kinetin was very poor. With stem-node explants under variable concentration of BAP, IPA-3088 performed best showing 17 buds per explants. Similarly Pusa-9 and IPA 3088 revea...
The Dof (DNA binding with one finger) transcription factor family represents a typical example of... more The Dof (DNA binding with one finger) transcription factor family represents a typical example of zinc finger transcription factors associated with diverse functions exclusively in plants. In the present study three dimensional structure of twenty one Dof proteins of sorghum were predicted by I-TASSER server based multiple threading method and the resulting modelled structures were validated by PROCHECK, Ramachandran Plot and QMEAN Z-score. These models were assigned PMDB IDs PM0079009 to PM0079021 and PM0079023 to PM0079030. The secondary structure revealed diversity in terms of sheets, beta-alpha-beta units, beta hairpin, beta bulges, strands, helices, helix-helix interactions, beta turns and gamma turns. The structural motif scan revealed presence of specific amino acid rich profile for each SbDof proteins along with a typical zinc finger Dof type profile. The putative functions were also predicted based on gene ontology (GO) terms in I-TASSER server. In-silico expression profili...
A total of 121 protein sequences of pectate lyases were subjected to homology search, multiple se... more A total of 121 protein sequences of pectate lyases were subjected to homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. The phylogenetic tree constructed revealed different clusters based on different source organisms representing bacterial, fungal, plant, and nematode pectate lyases. The multiple accessions of bacterial, fungal, nematode, and plant pectate lyase protein sequences were placed closely revealing a sequence level similarity. The multiple sequence alignment of these pectate lyase protein sequences from different source organisms showed conserved regions at different stretches with maximum homology from amino acid residues 439-467, 715-816, and 829-910 which could be used for designing degenerate primers or probes specific for pectate lyases. The motif analysis revealed a conserved Pec_Lyase_C domain uniformly observed in all pectate lyases irrespective of variable sources suggesting its possible role in structural and enzym...
An acidic polygalacturonase (PG) secreted by Rhizopus oryzae MTCC-1987 in submerged fermentation ... more An acidic polygalacturonase (PG) secreted by Rhizopus oryzae MTCC-1987 in submerged fermentation condition has been purified to electrophoretic homogeneity using ammonium sulphate fractionation and anion exchange chromatography on diethylaminoethyl cellulose. The purified enzyme gave a single protein band in sodium dodecyl sulphatepolyacrylamide gel electrophoresis analysis with a molecular mass corresponding to 75.5 kDa. The K m and k cat values of the PG were 2.7 mg/mL and 2.23 × 103 s−1, respectively, using citrus polygalacturonic acid as the substrate. The optimum pH of the purified PG was 5.0 and it does not loose activity appreciably if left for 24 hours in the pH range from 5.0 to 12.0. The optimum temperature of purified enzyme was 50°C and the enzyme does not loose activity below 30°C if exposed for two hours. The purified enzyme showed complete inhibition with 1 mM Ag+, Hg2+ and KMnO4, while it was stimulated to some extent by Co2+. The purified PG exhibited retting of Cro...
Genetic Engineering and Biotechnology Journal, 2010
The genetic relatedness of three varieties of finger millet (Eleusine coracana L.) with varying s... more The genetic relatedness of three varieties of finger millet (Eleusine coracana L.) with varying seed coat color namely PRM-1 (Brown), PRM-701 (White) and PRM-801 (Golden) were assessed with 10 RAPD and 10 ISSR markers. Molecular parameters viz., total number of bands, average polymorphic bands, average percent polymorphism, average polymorphic information content (PIC) and average expected gene diversity (Hi) generated from RAPD and ISSR markers has been compared. The RAPD profiling of ...
Physiology and molecular biology of plants : an international journal of functional plant biology, 2016
Nuclear factor Y (NF-Y) is a heterotrimeric transcription factor (TF) complex with preferential b... more Nuclear factor Y (NF-Y) is a heterotrimeric transcription factor (TF) complex with preferential binding to CCAAT elements of promoters, regulating gene expression in most of the higher eukaryotes. The availability of plant genome sequences have revealed multiple number of genes coding for the three subunits, namely NF-YA, NF-YB and NF-YC in contrast to single NF-Y gene for each subunit reported in yeast and animals. A total of 33 NF-YTF comprising of 8 NF-YA, 11 NF-YB and 14 NF-YC subunits were accessed from the sorghum genome. The bioinformatic characterization of NF-Y gene family of sorghum for gene structure, chromosome location, protein motif, phylogeny, gene duplication and in-silico expression under abiotic stresses have been attempted in the present study. The identified SbNF-Y genes are distributed on all the 10 chromosomes of sorghum with variability in the frequency and 18 out of 33 SbNF-Ys were found to be intronless. Segmental duplication event was found to be predominan...
An extracellular pectin lyase secreted by Fusarium decemcellulare MTCC 2079 under solid state fer... more An extracellular pectin lyase secreted by Fusarium decemcellulare MTCC 2079 under solid state fermentation condition has been purified to electrophoretic homogeniety by using ammonium sulfate fractionation, carboxymethyl cellulose and gel filtration (Sephadex G-100) column chromatographies. The purified enzyme showed single protein band corresponding to molecular mass 45 ± 01 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme had maximum activity at pH 9.0 and showed maximum stability in the pH range of 9.0–12.0. The optimum temperature of the purified enzyme was 50 °C and it showed maximum stability upto 40 °C. The energy of activation for the thermal denaturation (Ea) was 59.06 kJ mol−1 K−1. The Km and kcat values using citrus pectin as the substrate were 0.125 mg ml−1 and 72.9 s−1 in 100 mM sodium carbonate buffer pH 9.0 at 50 °C. The biophysical studies on pectin lyase showed that its secondary structure belongs to α + β class of protein with comparativ...
Heterotrimeric nuclear factors Y (NF-Ys) are involved in regulation of various vital functions in... more Heterotrimeric nuclear factors Y (NF-Ys) are involved in regulation of various vital functions in all eukaryotic organisms. Although a number of NF-Y subunits have been characterized in model plants, only a few have been functionally evaluated in crops. In this work, a number of genes encoding NF-YB and NF-YC subunits were isolated from drought-tolerant wheat (Triticum aestivum L. cv. RAC875), and the impact of the overexpression of TaNF-YB4 in the Australian wheat cultivar Gladius was investigated. TaNF-YB4 was isolated as a result of two consecutive yeast two-hybrid (Y2H) screens, where ZmNF-YB2a was used as a starting bait. A new NF-YC subunit, designated TaNF-YC15, was isolated in the first Y2H screen and used as bait in a second screen, which identified two wheat NF-YB subunits, TaNF-YB2 and TaNF-YB4. Three-dimensional modelling of a TaNF-YB2/TaNF-YC15 dimer revealed structural determinants that may underlie interaction selectivity. The TaNF-YB4 gene was placed under the contro...
Bacterial, fungal, plant and animal lipases amino acid sequences retrieved from different databas... more Bacterial, fungal, plant and animal lipases amino acid sequences retrieved from different databases were analyzed for homology search, multiple sequence alignment, phylogenetic and motif assessment using different bioinformatics tools. The multiple sequence alignment revealed different conserved stretches of amino acids exclusively for each groups along with a highly conserved glycine residue for plant and animal lipases. The phylogenetic tree constructed for different groups uniformly showed two major clusters divided into several sub-clusters based on the closeness of the lipases protein sequences irrespective of the source organisms. The 64 bacterial lipases protein sequences were distributed into two main clusters with various sub-clusters revealing the rich diversity of lipases among diverse bacterial strains. The fungal lipase protein sequences based phylogenetic tree revealed two major clusters for representing two important groups namely ascomycota and basidomycota with seve...
Microbial Diversity and Biotechnology in Food Security, 2014
The genomic sequences of Aspergillus flavus NRRL 3357 revealed seven pectin lyase (pnl) genes, wh... more The genomic sequences of Aspergillus flavus NRRL 3357 revealed seven pectin lyase (pnl) genes, which were used to design gene-specific primers and these genes were amplified through polymerase chain reaction (PCR) from five strains of A. flavus, namely A. flavus MTCC 7589, A. flavus MTCC 10938, A. flavus MTCC 8836, A. flavus NIICC 8142 and A. flavus NIICC 8147. All the seven predicted pnl genes were amplified from the genomic DNA of A. flavus NIICC 8142, while six pnl genes were from A. flavus MTCC8837 and five pnl genes from the remaining strains, respectively. A total of five pnl gene sequences of A. flavus NIICC 8142 designated as Afpnl-1, Afpnl-2, Afpnl-3, Afpnl-4, and Afpnl-5 were submitted to GenBank and assigned accession numbers JQ735890 to JQ735894, respectively. These sequences were subjected to in silico characterization for homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. The homology search revealed their identity to the predicted pectin lyase genes of A. flavus NRRL 3357, and multiple sequence alignment of these five genes showed various conserved residues. The phylogenetic tree revealed two distinct clusters and four subclusters with five pectin lyase genes of A. flavus occupying distinct position among the PCR-amplified pectin lyase genes from different fungi. The presence of unique pec_lyase C domain was observed among these sequences.
A neutral pectin lyase produced by a new fungal strain Oidiodendron echinulatum MTCC 1356 under s... more A neutral pectin lyase produced by a new fungal strain Oidiodendron echinulatum MTCC 1356 under solid state fermentation using wheat bran as agro waste has been studied. The enzyme was purified by ammonium sulphate precipitation (30-60%), DEAE anion exchange and Sephadex G-100 column chromatographies. The SDS-PAGE and native PAGE revealed two bands of sizes 42 and 47 kDa. The enzyme was purified 37 fold with specific activity of 4.5 U/mg and 2.25% yield. The K(m) and V(max) values determined using citrus pectin were 1.2 mg/ml and 0.36 IU/min respectively. The pH and temperature optima were pH 7.0 and 50 °C, respectively. The pH stability was around 5.0 for 24 h at 20 °C. The purified enzyme retained maximum activity for 30 min upto 50 °C. The activation energy for thermal denaturation of the purified enzyme was found to be 60.0 kJ/Mol. The effects of various metal ions and protein inhibitors on enzyme activity have revealed total inhibition of the enzyme activity in the presence of Ag(+) and Cu(+) and KMnO(4) at 1 mM. The neutral pectin lyase showed retting of Crotalaria juncea fibre in the presence of EDTA.
The diversity of Aspergillus from soil of Kusmi Forest, an Indian subtropical Teak forest at Gora... more The diversity of Aspergillus from soil of Kusmi Forest, an Indian subtropical Teak forest at Gorakhpur resulted in characterizing a total of 415 isolates which were then subjected to amylases production. The preliminary amylases production revealed a total of 14 promising isolates of Aspergillus comprising of ten different strains of A. flavus, two strains of A. tamarii, one strain each of A. niger and A. awamori which are characterized further. The optimum temperature of amylases produced from 14 isolates ranged from 30 ...
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