Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in sout... more Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in southern China and SouthEast Asia. The selection programme for growth trait has been conducted since 2010 in two selected lines for three generations, namely the Hainan (H) and Thailand (T) selected lines. Ten polymorphic microsatellite loci were used to monitor changes in their genetic diversity and structure during the selection process. A relatively high level of overall genetic diversity was observed, while the number of microsatellite allele declined slightly in the mass selection lines. Comparative diversity estimates indicated greater genetic diversity in H line than T line. Moreover, a relatively weak but significant genetic differentiation was found in both of the selected generations. In addition, the higher genetic variation observed in H line is most likely caused by artificial selection while selection bias on a growth advantage for the T population. These findings suggest no significant decline of genetic diversity was observed in the presently selected lines and that a potential exists for genetic improvement and development for B. areolata.
Journal of Molecular Graphics & Modelling, Sep 1, 2013
Insights into the structure-function relationship of disease resistance protein HCTR in maize (Ze... more Insights into the structure-function relationship of disease resistance protein HCTR in maize (Zea mays L.
The reproductive stage is the most important critical stage in rice where water is needed for the... more The reproductive stage is the most important critical stage in rice where water is needed for the formation of seeds. Water scarcity in this situation drastically reduces yield which in turn hampers farmers' effort towards agriculture. Traditional landraces do have resistant genes inside them against several abiotic and biotic stresses which can act as valuable donors to produce new gene combinations. A high-density SNP based genetic map was constructed by Genotyping by sequencing (GBS) technique using F7 recombinant inbred line (RIL) population developed from Banglami x Ranjit to identify tightly linked markers associated with QTLs. The total map length was 1306.424 cM which had an average inter-marker distance of 0.281 cM. A total of 42 QTLs were identi ed under both reproductive stage drought stress (RS) and non stress (NS) conditions for sixteen different yield and its component traits which explained a phenotypic variance (PVE) of 1.95-13.36 %. Four QTLs explained a PVE ≥ 10% which denotes they are major QTLs. Analysis of the genes present within the QTL regions revealed the presence of 6 candidate genes which were differentially expressed under stress condition. These genes and QTLs will be helpful for marker-assisted pyramiding to improve drought tolerance in rice. valuable material for developing drought tolerant varieties. Panda et al. (2021) reported that drought escape in rice through early owering and still giving good yield is an important trait which has to be considered while e ciently selecting drought tolerant genotypes.
Southeast Asia hosts a great diversity of different rattan genotypes. There are 5 genera and 60 d... more Southeast Asia hosts a great diversity of different rattan genotypes. There are 5 genera and 60 different species of rattan in India and the Northeastern region is a natural sanctuary for 4 different genera and 16 different species. The natural reserves of this species have come under the threat of genetic erosion due to overexploitation. This investigation was directed at characterizing 15 rattan genotypes of the genera Calamus, Plectocomia and Daemonorops which yield rattans of commercial importance, based on RAPD fingerprints. From 20 different random decamer primers, 12 primers gave reproducible amplification profiles and 104 polymorphic bands. A considerable degree of polymorphism (98.1%) was detected among the genotypes. Jaccard's coefficient of similarity ranged from 0.146 to 0.632 with a mean of 0.320-0.082, signifying extensive genetic divergence among the genotypes studied. UPGMA cluster analysis clearly distinguished P. assamica Griff. and C. erectus Roxb. The other 13 genotypes were grouped into two distinct clusters, one cluster involving two C. tenuis genotypes along with C. in-ermis T.Anders. and C. acanthospathus Griff. and the other cluster involving the rest of the Calamus genotypes along with D. jenkinsianus (Griff.) Mart. Unique fingerprints for 7 Calamus and 1 Daemonorops genotypes were detected. The results presented in this paper demonstrated the utility of RAPD markers in elucidating patterns of genetic variation among genotypes of the three main rattan genera of Northeast India and in identifying individual genotypes, which may serve as potential sources of unique genetic material for genetic improvement and conservation.
An in vitro approach became necessary in order to meet the increasing demand and conservation of ... more An in vitro approach became necessary in order to meet the increasing demand and conservation of the natural population of the economically important leguminous tree, Albizzia lebbeck. For this purpose, a study was undertaken to determine the amenability of apical buds from in vitro seedlings of A. lebbeck for direct shoot regeneration. The explants from 7-day-old seedlings were plated on Murashige and Skoog (MS) basal medium supplemented with 6benzylaminopurine (BAP) and kinetin solely or in combinations of both at different concentrations. The best response (68.33%) with an average of 3-7 shoots per explant and shoot length of 4.47 ± 0.49 cms was observed on MS basal medium supplemented with 1 mg/L 6benzylaminopurine. Kinetin was not found to be effective in induction of multiple shoot proliferation. In vitro rooting of the microshoots was tried in growth regulator-free as well as indole-3-acetic acid (IAA) or indole-3-butyric acid (IBA) fortified half strength MS medium. Highest rooting (55%) of the microshoots was achieved on half strength MS media supplemented with 1 mg/L indole-3-acetic acid (IAA) with an average of 2 shoots per shoot and shoot length of 3.87 ± 1.19 cms. Soilrite was found to be the best for acclimatization and establishment of the rooted plantlets, as compared to the other potting mixtures. The plantlets showed a survival percentage of 43.33%.
Biomedical Journal of Scientific & Technical Research, 2021
Functional genomics is the study of gene function and interaction with other proteins that contri... more Functional genomics is the study of gene function and interaction with other proteins that contribute to different metabolic pathways. The most commonly used approach for identifying gene function is to reduce or completely disrupt its normal expression pattern. Gene knock-down and gene knockout are two reverse genetic approaches that are widely used for discovering gene function. RNAi-mediated gene silencing has been used as a magic bullet over a decade to disrupt gene expression in many organisms. However, new genome editing tools-especially CRISPR-based technologies-has become major breakthrough against RNAi dominance in reverse genetics studies. RNAi results in temporary silencing of gene expression (knock-down) but CRISPR/Cas system results in permanent gene silencing (knock-out). Both of this loss of function technology has their own advantages and limitations. The goal of a biological experiment is the crucial determining factor for choosing the right tool. The whole article compares the mechanism, advantage and disadvantages of these two technologies.
Centella asiatica (L.) is one of the most valuable medicinal plants since prehistoric times. The ... more Centella asiatica (L.) is one of the most valuable medicinal plants since prehistoric times. The pharmaceutical importance of this herb is due to the accumulation of large quantities of pentacyclic triterpenoid saponins, collectively known as centelloids synthesized by the isoprenoid biosynthesis pathway. Biosynthesis of triterpenoid in the plants proceeds via either of the two pathways, viz. Mevalonate (MVA) pathway (in the cytosol) or 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway (in plastid). In Centella, the pathway leading to the accumulation of triterpenoid is still not known or elucidated. Thus, to know whether the MVA or MEP pathway or a cross-talk between the pathway leads to the biosynthesis of triterpenoid, silencing the key regulatory gene using RNAi tool, of each of the pathway and then analyze a metabolite is an efficient approach. The key regulatory enzyme of the MVA pathway i.e. 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) has already been successfully silenced using RNAi tool [1]. In the present study, the 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) a key regulatory enzyme in MEP pathway is silenced. The RNAi-DXR construct in pHANNIBAL vector was cloned into a binary vector pART27 and subsequently transformed into Agrobacterium strain AGL1. The transient analysis of the RNAi-CaDXR using semi-quantitative RT-PCR confirmed the silencing of the endogenous DXR gene in Nicotiana and further confirmed in Centella asiatica. The present study is the first step aimed to delineate the MEP pathway using RNAi silencing approach to elucidate its role in the accumulation of triterpenoid in this important medicinal plant.
A good quality RNA is an important prerequisite for any downstream molecular analysis. However, i... more A good quality RNA is an important prerequisite for any downstream molecular analysis. However, isolation of quality RNA from tea is difficult due to presence of high amount of phenols and polysaccharides. We describe here an efficient protocol for total RNA isolation without liquid nitrogen from apical buds and tender leaves of tea. This protocol yielded RNA 1248±23 μg/g of fresh leaf tissue with high purity. The RNA was also found to be suitable for mRNA purification, cDNA synthesis and cloning through RT-PCR (reverse transcription-polymerase chain reaction) and cDNA library construction.
International Journal of Current Microbiology and Applied Sciences, 2020
Molecular characterization of bumble bees has been carried out in the Department of Entomology, A... more Molecular characterization of bumble bees has been carried out in the Department of Entomology, Assam Agricultural University during the period 2015-2018. During the investigation, 5 (Five) species of bumble bees viz. Bombus orientalis Smith, B. buccinatoris Smith, B. tunicatus Smith, B. haemorrhoidalis Smith and B. miniatus Bingham have been recorded. Similarity matrix for Jaccard's Coefficient based on SSR banding of 5 bumble bee species ranged from 0.062 to 0.921 i.e. 6.2% to 92.1%. The dendrogram readily separated the bumble bee species into two main clusters (A and B). The cluster A includes Bombus orientalis and B. buccinatori while cluster B includes B. tunicatus, B. haemorrhoidalis and B. miniatus.
Amrit Sagar belonging to banana (Musa acuminata) genome group AAA is popularly grown in North Eas... more Amrit Sagar belonging to banana (Musa acuminata) genome group AAA is popularly grown in North Eastern part of India for its high yield and natural disease resistance potential. The explants were established initially on supplemented Murashige and Skoog's (MS) mediums followed by subculturing for multiple shoot induction. Various concentrations of BAP were tested to improve the quality and quantity of multiple shoots induction, out of which 10 mg/l BAP gave the best result. A total of 6-8 cycles of subcultures were carried out, each with an interval of 20-25 days. Among these, well established healthy shoots of 4-5 cm were transferred onto rooting medium containing 10 mg/l sucrose. After 3 weeks, plantlets were carefully acclimatized to adapt the green house condition and subsequently transferred to field. Appearance of off-types of plantlets during the Original Research Article
Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in sout... more Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in southern China and SouthEast Asia. The selection programme for growth trait has been conducted since 2010 in two selected lines for three generations, namely the Hainan (H) and Thailand (T) selected lines. Ten polymorphic microsatellite loci were used to monitor changes in their genetic diversity and structure during the selection process. A relatively high level of overall genetic diversity was observed, while the number of microsatellite allele declined slightly in the mass selection lines. Comparative diversity estimates indicated greater genetic diversity in H line than T line. Moreover, a relatively weak but significant genetic differentiation was found in both of the selected generations. In addition, the higher genetic variation observed in H line is most likely caused by artificial selection while selection bias on a growth advantage for the T population. These findings suggest no significant decline of genetic diversity was observed in the presently selected lines and that a potential exists for genetic improvement and development for B. areolata.
Citronella ( Cymbopogon winterianus) is one of the richest sources of high-value isoprenoid aroma... more Citronella ( Cymbopogon winterianus) is one of the richest sources of high-value isoprenoid aromatic compounds used as flavour, fragrance, and therapeutic elements. These isoprenoid compounds are synthesized by 2 independent pathways: mevalonate pathway and 2-C-methyl-d-erythritol-4-phosphate pathway. Evidence suggests that 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) is a rate-controlling enzyme for the synthesis of variety of isoprenoids. This study reports the isolation, characterization, and tissue-specific expression analysis of HMGR from citronella. The modelled HMGR is a class I type of HMGR enzyme with 3-domain architecture. The active site comprises a cofactor (nicotinamide adenine dinucleotide phosphate) and the substrate-binding motifs. The real-time and quantitative reverse transcription-polymerase chain reaction results revealed equal expression level in both leaf sheath and root tissue. The results from our study shall be a valuable resource for future molecular int...
The 3-hydroxy-3-methyl glutaryl CoA reductase (HMGR) is the key enzyme of mevalonate pathway in p... more The 3-hydroxy-3-methyl glutaryl CoA reductase (HMGR) is the key enzyme of mevalonate pathway in plants. A partial genomic DNA fragment encoding HMGR conserved domain (denoted as AbHMGR) is isolated from Aconitum balfourii Stapf. It comprises 871 bp encoding 290 amino acids. In silico analysis reveals that it had extensive similarities to other plant HMGR gene. Domain analysis of AbHMGR showed two highly conserved NADPH and HMG CoA domains. Docking study predicted inhibitor, substrate and cofactor binding sites in the protein. Expression analysis revealed that AbHMGR is similarly expressed in all tested tissues with differential pattern. The highest expression was found in leaf tissue. However, fold expression in root and shoot tissue was almost similar. Enzyme activity of HMGR was found to be much higher in leaf tissue as compared to other tissues. The highest aconitine content (0.015 %) was obtained in root tissues. Our data laid a foundation for further investigation of HMGR role in Aconitum balfourii. Keywords Aconitine Á Mevalonate pathway Á 3-Hydroxy-3-methylglutaryl coenzyme A reductase Á HMGR Á Expression profiling Abbreviations IPP Isopentyl pyrophosphate HMGR 3-Hydroxy-3-methyl glutaryl-CoA reductase MVA Mevalonate pathway MEP 2-C-methyl-D-erythritol 4-phosphate Electronic supplementary material The online version of this article (
The 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR; EC1.1.1.267), an NADPH-dependent reduct... more The 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR; EC1.1.1.267), an NADPH-dependent reductase, plays a pivotal role in the methylerythritol 4-phosphate pathway (MEP), in the conversion of 1-deoxy-D-xylulose-5-phosphate (DXP) into MEP. The sheath and leaf of citronella (Cymbopogon winterianus) accumulates large amount of terpenes and sesquiterpenes with proven medicinal value and economic uses. Thus, sequencing of full length dxr gene and its characterization seems to be a valuable resource in metabolic engineering to alter the flux of isoprenoid active ingredients in plants. In this study, full length DXR from citronella was characterized through in silico and tissue-specific expression studies to explain its structure-function mechanism, mode of cofactor recognition and differential expression. The modelled DXR has a three-domain architecture and its active site comprised of a cofactor (NADPH) binding pocket and the substrate-binding pocket. Molecular dynamics simulation studies indicated that DXR model retained most of its secondary structure during 10 ns simulation in aqueous solution. The modelled DXR superimposes well with its closest structural homolog but subtle variations in the charge distribution over the cofactor recognition site were noticed. Molecular docking study revealed critical residues aiding tight anchoring NADPH within the active pocket of DXR. Tissue-specific differential expression analysis using semi-quantitative RT-PCR and qRT-PCR in various tissues of citronella plant revealed distinct differential expression of DXR. To our knowledge, this is the first ever report on DXR from the important medicinal plant citronella and further characterization of this gene will open up better avenues for metabolic engineering of secondary metabolite pathway genes from medicinal plants in the near future.
Black gram (Vigna mungo) seeds are a rich source of digestible proteins, however, during storage ... more Black gram (Vigna mungo) seeds are a rich source of digestible proteins, however, during storage these seeds are severely damaged by bruchids (Callosobruchus spp.), reducing seed quality and yield losses. Most of the cultivated genotypes of black gram are susceptible to bruchids, however, few tolerant genotypes have also been identified but the mechanism of tolerance is poorly understood. We employed Suppression Subtractive Hybridization (SSH) to identify specifically, but rarely expressed bruchid egg induced genes in black gram. In this study, Suppression Subtractive Hybridization (SSH) library was constructed to study the genes involved in defense response in black gram against bruchid infestation. An EST library of 277 clones was obtained for further analyses. Based on CAP3 assembly, 134 unigenes were computationally annotated using Blast2GOPRO software. In all, 20 defense related genes were subject to quantitative PCR analysis (qPCR) out of which 12 genes showed up-regulation in...
The NADPH-dependent HC-toxin reductases (HCTR1 and 2) encoded by enzymatic class of disease resis... more The NADPH-dependent HC-toxin reductases (HCTR1 and 2) encoded by enzymatic class of disease resistance homologous genes (Hm1 and Hm2) protect maize by detoxifying a cyclic tetrapeptide, HC-toxin, secreted by the fungus Cochliobolus carbonum race 1(CCR1). Unlike the other classes' resistance (R) genes, HCTR-mediated disease resistance is an inimitable mechanism where the avirulence (Avr) component from CCR1 is not involved in toxin degradation. In this study, we attempted to decipher cofactor (NADPH) recognition and mode of HC-toxin binding to HCTRs through molecular docking, molecular dynamics (MD) simulations and binding free energy calculation methods. The rationality and the stability of docked complexes were validated by 30-ns MD simulation. The binding free energy decomposition of enzyme-cofactor complex was calculated to find the driving force behind cofactor recognition. The overall binding free energies of HCTR1-NADPH and HCTR2-NADPH were found to be 2616.989 and 216.9749 kJ mol 21 respectively. The binding free energy decomposition revealed that the binding of NADPH to the HCTR1 is mainly governed by van der Waals and nonpolar interactions, whereas electrostatic terms play dominant role in stabilizing the binding mode between HCTR2 and NADPH. Further, docking analysis of HC-toxin with HCTR-NADPH complexes showed a distinct mode of binding and the complexes were stabilized by a strong network of hydrogen bond and hydrophobic interactions. This study is the first in silico attempt to unravel the biophysical and biochemical basis of cofactor recognition in enzymatic class of R genes in cereal crop maize. Citation: Dehury B, Patra MC, Maharana J, Sahu J, Sen P, et al. (2014) Structure-Based Computational Study of Two Disease Resistance Gene Homologues (Hm1 and Hm2) in Maize (Zea mays L.) with Implications in Plant-Pathogen Interactions. PLoS ONE 9(5): e97852.
Glutathione synthetase (gshB) has previously been reported to confer tolerance to acidic soil con... more Glutathione synthetase (gshB) has previously been reported to confer tolerance to acidic soil condition in Rhizobium species. Cloning the gene coding for this enzyme necessitates the designing of proper primer sets which in turn depends on the identification of high quality sequence similarity in multiple global alignments. In this experiment, a group of homologous gene sequences related to gshB gene (accession no: gi-86355669:327589-328536) of Rhizobium etli CFN 42, were extracted from NCBI nucleotide sequence databases using BLASTN and were analyzed for designing degenerate primers. However, the T-coffee multiple global alignment results did not show any block of conserved region for the above sequence set to design the primers. Therefore, we attempted to identify the location of common motif region based on multiple local alignments employing the MEME algorithm supported with MAST and Primer3. The results revealed some common motif regions that enabled us to design the primer sets for related gshB gene sequences. The result will be validated in wet lab.
Journal of Plant Biochemistry and Biotechnology, 2013
Superoxide dismutases (SODs), members of the metalloenzymes family are most effective intracellul... more Superoxide dismutases (SODs), members of the metalloenzymes family are most effective intracellular enzymatic antioxidant in aerobic organisms. These enzymes provide the first line of defense in plants against the toxic effects of elevated levels of reactive oxygen species (ROS) generated during various environmental stresses. The availability of high-throughput computational tools has provided better opportunities to characterize the protein features and determine their function. In the present study an attempt was made to gain an insight into the structure and evolution of subunits of SODs (Cu-Zn, Mn and Fe SODs) of rice. The 3-Dimensional structures of SODs were modeled based on available X-ray crystal structures and further validated. The primary sequence, secondary and tertiary structure analysis revealed Mn and Fe SOD to be structurally homologous while Cu-Zn SOD is un-related to either of them. Comparative structural study also revealed former two were dominated by α-helices followed by β-strands in contrast; Cu-Zn SOD dominated by β-strands.
Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in sout... more Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in southern China and SouthEast Asia. The selection programme for growth trait has been conducted since 2010 in two selected lines for three generations, namely the Hainan (H) and Thailand (T) selected lines. Ten polymorphic microsatellite loci were used to monitor changes in their genetic diversity and structure during the selection process. A relatively high level of overall genetic diversity was observed, while the number of microsatellite allele declined slightly in the mass selection lines. Comparative diversity estimates indicated greater genetic diversity in H line than T line. Moreover, a relatively weak but significant genetic differentiation was found in both of the selected generations. In addition, the higher genetic variation observed in H line is most likely caused by artificial selection while selection bias on a growth advantage for the T population. These findings suggest no significant decline of genetic diversity was observed in the presently selected lines and that a potential exists for genetic improvement and development for B. areolata.
Journal of Molecular Graphics & Modelling, Sep 1, 2013
Insights into the structure-function relationship of disease resistance protein HCTR in maize (Ze... more Insights into the structure-function relationship of disease resistance protein HCTR in maize (Zea mays L.
The reproductive stage is the most important critical stage in rice where water is needed for the... more The reproductive stage is the most important critical stage in rice where water is needed for the formation of seeds. Water scarcity in this situation drastically reduces yield which in turn hampers farmers' effort towards agriculture. Traditional landraces do have resistant genes inside them against several abiotic and biotic stresses which can act as valuable donors to produce new gene combinations. A high-density SNP based genetic map was constructed by Genotyping by sequencing (GBS) technique using F7 recombinant inbred line (RIL) population developed from Banglami x Ranjit to identify tightly linked markers associated with QTLs. The total map length was 1306.424 cM which had an average inter-marker distance of 0.281 cM. A total of 42 QTLs were identi ed under both reproductive stage drought stress (RS) and non stress (NS) conditions for sixteen different yield and its component traits which explained a phenotypic variance (PVE) of 1.95-13.36 %. Four QTLs explained a PVE ≥ 10% which denotes they are major QTLs. Analysis of the genes present within the QTL regions revealed the presence of 6 candidate genes which were differentially expressed under stress condition. These genes and QTLs will be helpful for marker-assisted pyramiding to improve drought tolerance in rice. valuable material for developing drought tolerant varieties. Panda et al. (2021) reported that drought escape in rice through early owering and still giving good yield is an important trait which has to be considered while e ciently selecting drought tolerant genotypes.
Southeast Asia hosts a great diversity of different rattan genotypes. There are 5 genera and 60 d... more Southeast Asia hosts a great diversity of different rattan genotypes. There are 5 genera and 60 different species of rattan in India and the Northeastern region is a natural sanctuary for 4 different genera and 16 different species. The natural reserves of this species have come under the threat of genetic erosion due to overexploitation. This investigation was directed at characterizing 15 rattan genotypes of the genera Calamus, Plectocomia and Daemonorops which yield rattans of commercial importance, based on RAPD fingerprints. From 20 different random decamer primers, 12 primers gave reproducible amplification profiles and 104 polymorphic bands. A considerable degree of polymorphism (98.1%) was detected among the genotypes. Jaccard's coefficient of similarity ranged from 0.146 to 0.632 with a mean of 0.320-0.082, signifying extensive genetic divergence among the genotypes studied. UPGMA cluster analysis clearly distinguished P. assamica Griff. and C. erectus Roxb. The other 13 genotypes were grouped into two distinct clusters, one cluster involving two C. tenuis genotypes along with C. in-ermis T.Anders. and C. acanthospathus Griff. and the other cluster involving the rest of the Calamus genotypes along with D. jenkinsianus (Griff.) Mart. Unique fingerprints for 7 Calamus and 1 Daemonorops genotypes were detected. The results presented in this paper demonstrated the utility of RAPD markers in elucidating patterns of genetic variation among genotypes of the three main rattan genera of Northeast India and in identifying individual genotypes, which may serve as potential sources of unique genetic material for genetic improvement and conservation.
An in vitro approach became necessary in order to meet the increasing demand and conservation of ... more An in vitro approach became necessary in order to meet the increasing demand and conservation of the natural population of the economically important leguminous tree, Albizzia lebbeck. For this purpose, a study was undertaken to determine the amenability of apical buds from in vitro seedlings of A. lebbeck for direct shoot regeneration. The explants from 7-day-old seedlings were plated on Murashige and Skoog (MS) basal medium supplemented with 6benzylaminopurine (BAP) and kinetin solely or in combinations of both at different concentrations. The best response (68.33%) with an average of 3-7 shoots per explant and shoot length of 4.47 ± 0.49 cms was observed on MS basal medium supplemented with 1 mg/L 6benzylaminopurine. Kinetin was not found to be effective in induction of multiple shoot proliferation. In vitro rooting of the microshoots was tried in growth regulator-free as well as indole-3-acetic acid (IAA) or indole-3-butyric acid (IBA) fortified half strength MS medium. Highest rooting (55%) of the microshoots was achieved on half strength MS media supplemented with 1 mg/L indole-3-acetic acid (IAA) with an average of 2 shoots per shoot and shoot length of 3.87 ± 1.19 cms. Soilrite was found to be the best for acclimatization and establishment of the rooted plantlets, as compared to the other potting mixtures. The plantlets showed a survival percentage of 43.33%.
Biomedical Journal of Scientific & Technical Research, 2021
Functional genomics is the study of gene function and interaction with other proteins that contri... more Functional genomics is the study of gene function and interaction with other proteins that contribute to different metabolic pathways. The most commonly used approach for identifying gene function is to reduce or completely disrupt its normal expression pattern. Gene knock-down and gene knockout are two reverse genetic approaches that are widely used for discovering gene function. RNAi-mediated gene silencing has been used as a magic bullet over a decade to disrupt gene expression in many organisms. However, new genome editing tools-especially CRISPR-based technologies-has become major breakthrough against RNAi dominance in reverse genetics studies. RNAi results in temporary silencing of gene expression (knock-down) but CRISPR/Cas system results in permanent gene silencing (knock-out). Both of this loss of function technology has their own advantages and limitations. The goal of a biological experiment is the crucial determining factor for choosing the right tool. The whole article compares the mechanism, advantage and disadvantages of these two technologies.
Centella asiatica (L.) is one of the most valuable medicinal plants since prehistoric times. The ... more Centella asiatica (L.) is one of the most valuable medicinal plants since prehistoric times. The pharmaceutical importance of this herb is due to the accumulation of large quantities of pentacyclic triterpenoid saponins, collectively known as centelloids synthesized by the isoprenoid biosynthesis pathway. Biosynthesis of triterpenoid in the plants proceeds via either of the two pathways, viz. Mevalonate (MVA) pathway (in the cytosol) or 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway (in plastid). In Centella, the pathway leading to the accumulation of triterpenoid is still not known or elucidated. Thus, to know whether the MVA or MEP pathway or a cross-talk between the pathway leads to the biosynthesis of triterpenoid, silencing the key regulatory gene using RNAi tool, of each of the pathway and then analyze a metabolite is an efficient approach. The key regulatory enzyme of the MVA pathway i.e. 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) has already been successfully silenced using RNAi tool [1]. In the present study, the 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) a key regulatory enzyme in MEP pathway is silenced. The RNAi-DXR construct in pHANNIBAL vector was cloned into a binary vector pART27 and subsequently transformed into Agrobacterium strain AGL1. The transient analysis of the RNAi-CaDXR using semi-quantitative RT-PCR confirmed the silencing of the endogenous DXR gene in Nicotiana and further confirmed in Centella asiatica. The present study is the first step aimed to delineate the MEP pathway using RNAi silencing approach to elucidate its role in the accumulation of triterpenoid in this important medicinal plant.
A good quality RNA is an important prerequisite for any downstream molecular analysis. However, i... more A good quality RNA is an important prerequisite for any downstream molecular analysis. However, isolation of quality RNA from tea is difficult due to presence of high amount of phenols and polysaccharides. We describe here an efficient protocol for total RNA isolation without liquid nitrogen from apical buds and tender leaves of tea. This protocol yielded RNA 1248±23 μg/g of fresh leaf tissue with high purity. The RNA was also found to be suitable for mRNA purification, cDNA synthesis and cloning through RT-PCR (reverse transcription-polymerase chain reaction) and cDNA library construction.
International Journal of Current Microbiology and Applied Sciences, 2020
Molecular characterization of bumble bees has been carried out in the Department of Entomology, A... more Molecular characterization of bumble bees has been carried out in the Department of Entomology, Assam Agricultural University during the period 2015-2018. During the investigation, 5 (Five) species of bumble bees viz. Bombus orientalis Smith, B. buccinatoris Smith, B. tunicatus Smith, B. haemorrhoidalis Smith and B. miniatus Bingham have been recorded. Similarity matrix for Jaccard's Coefficient based on SSR banding of 5 bumble bee species ranged from 0.062 to 0.921 i.e. 6.2% to 92.1%. The dendrogram readily separated the bumble bee species into two main clusters (A and B). The cluster A includes Bombus orientalis and B. buccinatori while cluster B includes B. tunicatus, B. haemorrhoidalis and B. miniatus.
Amrit Sagar belonging to banana (Musa acuminata) genome group AAA is popularly grown in North Eas... more Amrit Sagar belonging to banana (Musa acuminata) genome group AAA is popularly grown in North Eastern part of India for its high yield and natural disease resistance potential. The explants were established initially on supplemented Murashige and Skoog's (MS) mediums followed by subculturing for multiple shoot induction. Various concentrations of BAP were tested to improve the quality and quantity of multiple shoots induction, out of which 10 mg/l BAP gave the best result. A total of 6-8 cycles of subcultures were carried out, each with an interval of 20-25 days. Among these, well established healthy shoots of 4-5 cm were transferred onto rooting medium containing 10 mg/l sucrose. After 3 weeks, plantlets were carefully acclimatized to adapt the green house condition and subsequently transferred to field. Appearance of off-types of plantlets during the Original Research Article
Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in sout... more Ivory shell, Babylonia areolata, is one of the commercially important mariculture species in southern China and SouthEast Asia. The selection programme for growth trait has been conducted since 2010 in two selected lines for three generations, namely the Hainan (H) and Thailand (T) selected lines. Ten polymorphic microsatellite loci were used to monitor changes in their genetic diversity and structure during the selection process. A relatively high level of overall genetic diversity was observed, while the number of microsatellite allele declined slightly in the mass selection lines. Comparative diversity estimates indicated greater genetic diversity in H line than T line. Moreover, a relatively weak but significant genetic differentiation was found in both of the selected generations. In addition, the higher genetic variation observed in H line is most likely caused by artificial selection while selection bias on a growth advantage for the T population. These findings suggest no significant decline of genetic diversity was observed in the presently selected lines and that a potential exists for genetic improvement and development for B. areolata.
Citronella ( Cymbopogon winterianus) is one of the richest sources of high-value isoprenoid aroma... more Citronella ( Cymbopogon winterianus) is one of the richest sources of high-value isoprenoid aromatic compounds used as flavour, fragrance, and therapeutic elements. These isoprenoid compounds are synthesized by 2 independent pathways: mevalonate pathway and 2-C-methyl-d-erythritol-4-phosphate pathway. Evidence suggests that 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) is a rate-controlling enzyme for the synthesis of variety of isoprenoids. This study reports the isolation, characterization, and tissue-specific expression analysis of HMGR from citronella. The modelled HMGR is a class I type of HMGR enzyme with 3-domain architecture. The active site comprises a cofactor (nicotinamide adenine dinucleotide phosphate) and the substrate-binding motifs. The real-time and quantitative reverse transcription-polymerase chain reaction results revealed equal expression level in both leaf sheath and root tissue. The results from our study shall be a valuable resource for future molecular int...
The 3-hydroxy-3-methyl glutaryl CoA reductase (HMGR) is the key enzyme of mevalonate pathway in p... more The 3-hydroxy-3-methyl glutaryl CoA reductase (HMGR) is the key enzyme of mevalonate pathway in plants. A partial genomic DNA fragment encoding HMGR conserved domain (denoted as AbHMGR) is isolated from Aconitum balfourii Stapf. It comprises 871 bp encoding 290 amino acids. In silico analysis reveals that it had extensive similarities to other plant HMGR gene. Domain analysis of AbHMGR showed two highly conserved NADPH and HMG CoA domains. Docking study predicted inhibitor, substrate and cofactor binding sites in the protein. Expression analysis revealed that AbHMGR is similarly expressed in all tested tissues with differential pattern. The highest expression was found in leaf tissue. However, fold expression in root and shoot tissue was almost similar. Enzyme activity of HMGR was found to be much higher in leaf tissue as compared to other tissues. The highest aconitine content (0.015 %) was obtained in root tissues. Our data laid a foundation for further investigation of HMGR role in Aconitum balfourii. Keywords Aconitine Á Mevalonate pathway Á 3-Hydroxy-3-methylglutaryl coenzyme A reductase Á HMGR Á Expression profiling Abbreviations IPP Isopentyl pyrophosphate HMGR 3-Hydroxy-3-methyl glutaryl-CoA reductase MVA Mevalonate pathway MEP 2-C-methyl-D-erythritol 4-phosphate Electronic supplementary material The online version of this article (
The 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR; EC1.1.1.267), an NADPH-dependent reduct... more The 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR; EC1.1.1.267), an NADPH-dependent reductase, plays a pivotal role in the methylerythritol 4-phosphate pathway (MEP), in the conversion of 1-deoxy-D-xylulose-5-phosphate (DXP) into MEP. The sheath and leaf of citronella (Cymbopogon winterianus) accumulates large amount of terpenes and sesquiterpenes with proven medicinal value and economic uses. Thus, sequencing of full length dxr gene and its characterization seems to be a valuable resource in metabolic engineering to alter the flux of isoprenoid active ingredients in plants. In this study, full length DXR from citronella was characterized through in silico and tissue-specific expression studies to explain its structure-function mechanism, mode of cofactor recognition and differential expression. The modelled DXR has a three-domain architecture and its active site comprised of a cofactor (NADPH) binding pocket and the substrate-binding pocket. Molecular dynamics simulation studies indicated that DXR model retained most of its secondary structure during 10 ns simulation in aqueous solution. The modelled DXR superimposes well with its closest structural homolog but subtle variations in the charge distribution over the cofactor recognition site were noticed. Molecular docking study revealed critical residues aiding tight anchoring NADPH within the active pocket of DXR. Tissue-specific differential expression analysis using semi-quantitative RT-PCR and qRT-PCR in various tissues of citronella plant revealed distinct differential expression of DXR. To our knowledge, this is the first ever report on DXR from the important medicinal plant citronella and further characterization of this gene will open up better avenues for metabolic engineering of secondary metabolite pathway genes from medicinal plants in the near future.
Black gram (Vigna mungo) seeds are a rich source of digestible proteins, however, during storage ... more Black gram (Vigna mungo) seeds are a rich source of digestible proteins, however, during storage these seeds are severely damaged by bruchids (Callosobruchus spp.), reducing seed quality and yield losses. Most of the cultivated genotypes of black gram are susceptible to bruchids, however, few tolerant genotypes have also been identified but the mechanism of tolerance is poorly understood. We employed Suppression Subtractive Hybridization (SSH) to identify specifically, but rarely expressed bruchid egg induced genes in black gram. In this study, Suppression Subtractive Hybridization (SSH) library was constructed to study the genes involved in defense response in black gram against bruchid infestation. An EST library of 277 clones was obtained for further analyses. Based on CAP3 assembly, 134 unigenes were computationally annotated using Blast2GOPRO software. In all, 20 defense related genes were subject to quantitative PCR analysis (qPCR) out of which 12 genes showed up-regulation in...
The NADPH-dependent HC-toxin reductases (HCTR1 and 2) encoded by enzymatic class of disease resis... more The NADPH-dependent HC-toxin reductases (HCTR1 and 2) encoded by enzymatic class of disease resistance homologous genes (Hm1 and Hm2) protect maize by detoxifying a cyclic tetrapeptide, HC-toxin, secreted by the fungus Cochliobolus carbonum race 1(CCR1). Unlike the other classes' resistance (R) genes, HCTR-mediated disease resistance is an inimitable mechanism where the avirulence (Avr) component from CCR1 is not involved in toxin degradation. In this study, we attempted to decipher cofactor (NADPH) recognition and mode of HC-toxin binding to HCTRs through molecular docking, molecular dynamics (MD) simulations and binding free energy calculation methods. The rationality and the stability of docked complexes were validated by 30-ns MD simulation. The binding free energy decomposition of enzyme-cofactor complex was calculated to find the driving force behind cofactor recognition. The overall binding free energies of HCTR1-NADPH and HCTR2-NADPH were found to be 2616.989 and 216.9749 kJ mol 21 respectively. The binding free energy decomposition revealed that the binding of NADPH to the HCTR1 is mainly governed by van der Waals and nonpolar interactions, whereas electrostatic terms play dominant role in stabilizing the binding mode between HCTR2 and NADPH. Further, docking analysis of HC-toxin with HCTR-NADPH complexes showed a distinct mode of binding and the complexes were stabilized by a strong network of hydrogen bond and hydrophobic interactions. This study is the first in silico attempt to unravel the biophysical and biochemical basis of cofactor recognition in enzymatic class of R genes in cereal crop maize. Citation: Dehury B, Patra MC, Maharana J, Sahu J, Sen P, et al. (2014) Structure-Based Computational Study of Two Disease Resistance Gene Homologues (Hm1 and Hm2) in Maize (Zea mays L.) with Implications in Plant-Pathogen Interactions. PLoS ONE 9(5): e97852.
Glutathione synthetase (gshB) has previously been reported to confer tolerance to acidic soil con... more Glutathione synthetase (gshB) has previously been reported to confer tolerance to acidic soil condition in Rhizobium species. Cloning the gene coding for this enzyme necessitates the designing of proper primer sets which in turn depends on the identification of high quality sequence similarity in multiple global alignments. In this experiment, a group of homologous gene sequences related to gshB gene (accession no: gi-86355669:327589-328536) of Rhizobium etli CFN 42, were extracted from NCBI nucleotide sequence databases using BLASTN and were analyzed for designing degenerate primers. However, the T-coffee multiple global alignment results did not show any block of conserved region for the above sequence set to design the primers. Therefore, we attempted to identify the location of common motif region based on multiple local alignments employing the MEME algorithm supported with MAST and Primer3. The results revealed some common motif regions that enabled us to design the primer sets for related gshB gene sequences. The result will be validated in wet lab.
Journal of Plant Biochemistry and Biotechnology, 2013
Superoxide dismutases (SODs), members of the metalloenzymes family are most effective intracellul... more Superoxide dismutases (SODs), members of the metalloenzymes family are most effective intracellular enzymatic antioxidant in aerobic organisms. These enzymes provide the first line of defense in plants against the toxic effects of elevated levels of reactive oxygen species (ROS) generated during various environmental stresses. The availability of high-throughput computational tools has provided better opportunities to characterize the protein features and determine their function. In the present study an attempt was made to gain an insight into the structure and evolution of subunits of SODs (Cu-Zn, Mn and Fe SODs) of rice. The 3-Dimensional structures of SODs were modeled based on available X-ray crystal structures and further validated. The primary sequence, secondary and tertiary structure analysis revealed Mn and Fe SOD to be structurally homologous while Cu-Zn SOD is un-related to either of them. Comparative structural study also revealed former two were dominated by α-helices followed by β-strands in contrast; Cu-Zn SOD dominated by β-strands.
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Papers by Priyabrata Sen