Papers by Selvaraju Kanagarajan
Sensors and Actuators B: Chemical, Feb 29, 2024
Agronomy, Sep 17, 2023
Melatonin (MT), a naturally occurring compound, is found in various species worldwide. In 1958, i... more Melatonin (MT), a naturally occurring compound, is found in various species worldwide. In 1958, it was first identified in the pineal gland of dairy cows. MT is an "old friend" but a "new compound" for plant biology. It brings experts and research minds from the broad field of plant sciences due to its considerable influence on plant systems. The MT production process in plants and animals is distinct, where it has been expressed explicitly in chloroplasts and mitochondria in plants. Tryptophan acts as the precursor for the formation of phyto-melatonin, along with intermediates including tryptamine, serotonin, N-acetyl serotonin, and 5-methoxy tryptamine. It plays a vital role in growth phases such as the seed germination and seedling growth of crop plants. MT significantly impacts the gas exchange, thereby improving physio-chemical functions in plant systems. During stress, the excessive generation and accumulation of reactive oxygen species (ROS) causes protein oxidation, lipid peroxidation, nucleic acid damage, and enzyme inhibition. Because it directly acts as an antioxidant compound, it awakens the plant antioxidant defense system during stress and reduces the production of ROS, which results in decreasing cellular oxidative damage. MT can enhance plant growth and development in response to various abiotic stresses such as drought, salinity, high temperature, flooding, and heavy metals by regulating the antioxidant mechanism of plants. However, these reactions differ significantly from crop to crop and are based on the level and kind of stress. The role of MT in the physiological functions of plants towards plant growth and development, tolerance towards various abiotic stresses, and approaches for enhancing the endogenous MT in plant systems are broadly reviewed and it is suggested that MT is a steering compound in directing major physiological functions of plants under the changing climate in future.
Genes, Sep 2, 2023
Ethiopian mustard (Brassica carinata A. Braun) is currently one of the potential oilseeds dedicat... more Ethiopian mustard (Brassica carinata A. Braun) is currently one of the potential oilseeds dedicated to the production for biofuel and other bio-industrial applications. The crop is assumed to be native to Ethiopia where a number of diversified B. carinata germplasms are found and conserved ex situ. However, there is very limited information on the genetic diversity and population structure of the species. This study aimed to investigate the genetic diversity and population structure of B. carinata genotypes of different origens using high-throughput single nucleotide polymorphism (SNP) markers. We used Brassica 90K Illumina Infinium TM SNP array for genotyping 90 B. carinata genotypes, and a total of 11,499 informative SNP markers were used for investigating the population structure and genetic diversity. The structure analysis, principal coordinate analysis (PcoA) and neighbor-joining tree analysis clustered the 90 B. carinata genotypes into two distinct subpopulations (Pop1 and Pop2). The majority of accessions (65%) were clustered in Pop1, mainly obtained from Oromia and South West Ethiopian People (SWEP) regions. Pop2 constituted dominantly of breeding lines and varieties, implying target selection contributed to the formation of distinct populations. Analysis of molecular variance (AMOVA) revealed a higher genetic variation (93%) within populations than between populations (7%), with low genetic differentiation (PhiPT = 0.07) and poor correlation between genetic and geographical distance (R = 0.02). This implies the presence of gene flow (Nm > 1) and weak geographical structure of accessions. Genetic diversity indices showed the presence of moderate genetic diversity in B. carinata populations with an average genetic diversity value (HE = 0.31) and polymorphism information content (PIC = 0.26). The findings of this study provide important and relevant information for future breeding and conservation efforts of B. carinata.
Current Issues in Molecular Biology
Plant metabolomics is a rapidly advancing field of plant sciences and systems biology. It involve... more Plant metabolomics is a rapidly advancing field of plant sciences and systems biology. It involves comprehensive analyses of small molecules (metabolites) in plant tissues and cells. These metabolites include a wide range of compounds, such as sugars, amino acids, organic acids, secondary metabolites (e.g., alkaloids and flavonoids), lipids, and more. Metabolomics allows an understanding of the functional roles of specific metabolites in plants’ physiology, development, and responses to biotic and abiotic stresses. It can lead to the identification of metabolites linked with specific traits or functions. Plant metabolic networks and pathways can be better understood with the help of metabolomics. Researchers can determine how plants react to environmental cues or genetic modifications by examining how metabolite profiles change under various crop stages. Metabolomics plays a major role in crop improvement and biotechnology. Integrating metabolomics data with other omics data (genomi...
PLOS ONE, 2012
The influenza A virus is of global concern for the poultry industry, especially the H5 and H7 sub... more The influenza A virus is of global concern for the poultry industry, especially the H5 and H7 subtypes as they have the potential to become highly pathogenic for poultry. In this study, the hemagglutinin (HA) of a low pathogenic avian influenza virus of the H7N7 subtype isolated from a Swedish mallard Anas platyrhynchos was sequenced, characterized and transiently expressed in Nicotiana benthamiana. Recently, plant expression systems have gained interest as an alternative for the production of vaccine antigens. To examine the possibility of expressing the HA protein in N. benthamiana, a cDNA fragment encoding the HA gene was synthesized de novo, modified with a Kozak sequence, a PR1a signal peptide, a Cterminal hexahistidine (66His) tag, and an endoplasmic retention signal (SEKDEL). The construct was cloned into a Cowpea mosaic virus (CPMV)-based vector (pEAQ-HT) and the resulting pEAQ-HT-HA plasmid, along with a vector (pJL3:p19) containing the viral gene-silencing suppressor p19 from Tomato bushy stunt virus, was agro-infiltrated into N. benthamiana. The highest gene expression of recombinant plant-produced, uncleaved HA (rHA0), as measured by quantitative real-time PCR was detected at 6 days post infiltration (dpi). Guided by the gene expression profile, rHA0 protein was extracted at 6 dpi and subsequently purified utilizing the 66His tag and immobilized metal ion adsorption chromatography. The yield was 0.2 g purified protein per kg fresh weight of leaves. Further molecular characterizations showed that the purified rHA0 protein was N-glycosylated and its identity confirmed by liquid chromatography-tandem mass spectrometry. In addition, the purified rHA0 exhibited hemagglutination and hemagglutination inhibition activity indicating that the rHA0 shares structural and functional properties with native HA protein of H7 influenza virus. Our results indicate that rHA0 maintained its native antigenicity and specificity, providing a good source of vaccine antigen to induce immune response in poultry species.
Frontiers in Plant Science, Jan 9, 2017
The wild species field cress (Lepidium campestre), belonging to the Brassicaceae family, has pote... more The wild species field cress (Lepidium campestre), belonging to the Brassicaceae family, has potential to be developed into a novel oilseed-and catch crop, however, the species needs to be further improved regarding some important agronomic traits. One of them is its low oil content which needs to be increased. As far as we know there is no study aiming at increasing the oil content that has been reported in this species. In order to investigate the possibility to increase the seed oil content in field cress, we have tried to introduce the Arabidopsis WRINKLED1 (AtWRI1) or hemoglobin (Hb) genes from either Arabidopsis thaliana (AtHb2) or Beta vulgaris (BvHb2) into field cress with the seed specific expression. The hypothesis was that the oil content would be increased by overexpressing these target genes. The results showed that the oil content was indeed increased by up to 29.9, 20.2, and 25.9% in the transgenic lines expressing AtWRI1, AtHb2, and BvHb2, respectively. The seed oil composition of the transgenic lines did not significantly deviate from the seed oil composition of the wild type plants. Our results indicate that genetic modification can be used in this wild species for its fast domestication into a future economically viable oilseed and catch crop.
Molecular Plant, Jun 1, 2016
was awarded the 2015 Nobel Prize in Physiology or Medicine for ''her discoveries concerning a nov... more was awarded the 2015 Nobel Prize in Physiology or Medicine for ''her discoveries concerning a novel therapy against malaria'' (). This therapy is based on the sesquiterpenoid artemisinin, isolated from Artemisia annua. Artemisinin and its derivatives, administered in the form of artemisinin-based combination therapies (ACTs), are currently the best treatment for malaria. We thank professor Kexuan Tang, Shanghai Jiao Tong University, for seeds of A. annua var. Chongqing. No conflict of interest declared.
Plant Molecular Biology, Nov 19, 2012
Artemisia annua L. produces small amounts of the sesquiterpenoid artemisinin, which is used for t... more Artemisia annua L. produces small amounts of the sesquiterpenoid artemisinin, which is used for treatment of malaria. A worldwide shortage of the drug has led to intense research to increase the yield of artemisinin in the plant. In order to study the regulation of expression of a key enzyme of artemisinin biosynthesis, the promoter region of the key enzyme amorpha-4,11-diene synthase (ADS) was cloned and fused with the β-glucuronidase (GUS) reporter gene. Transgenic plants of A. annua expressing this fusion were generated and studied. Transgenic plants expressing the GUS gene were used to establish the activity of the cloned promoter by a GUS activity staining procedure. GUS under the control of the ADS promoter showed specific expression in glandular trichomes. The activity of the ADS promoter varies temporally and in old tissues essentially no GUS staining could be observed. The expression pattern of GUS and ADS in aerial parts of the transgenic plant was essentially the same indicating that the cis-elements controlling glandular trichome specific expression are included in the cloned promoter. However, some cis-element(s) that control expression in root and old leaf appears to be missing in the cloned promoter. Furthermore, qPCR was used to compare the activity of the wild-type ADS promoter with that of the cloned ADS promoter. The latter promoter showed a considerably lower activity than the wild-type promoter as judged from the levels of GUS and ADS transcripts, respectively, which may be due to the removal of an enhancing cis-element from the ADS promoter. The ADS gene is specifically expressed in stalk and secretory cells of glandular trichomes of A. annua.
Frontiers in Plant Science, Mar 22, 2023
Improved fatty acid composition of field cress (Lepidium campestre) by CRISPR/Cas9-mediated genom... more Improved fatty acid composition of field cress (Lepidium campestre) by CRISPR/Cas9-mediated genome editing
Frontiers in Plant Science, Mar 22, 2023
Efficient protoplast regeneration protocol and CRISPR/Cas9-mediated editing of glucosinolate tran... more Efficient protoplast regeneration protocol and CRISPR/Cas9-mediated editing of glucosinolate transporter (GTR) genes in rapeseed (Brassica napus L.
Agronomy, Mar 26, 2022
Horse gram is one of the lesser-known beans widely grown in India. One hundred and twenty-three h... more Horse gram is one of the lesser-known beans widely grown in India. One hundred and twenty-three homozygous horse gram mutants were screened for powdery mildew (PM) disease resistance using the grade 0 to 4. The mutants were grouped based on the disease level of 0 to 2 (resistant) and susceptible (3 to 4). The PM altered the chlorophyll fluorescence (a/b ratio), maturity duration, and yield attributing traits. The yield loss ranged from 4.55% to 72.66%. After affirming the resistance level, the resistant mutant (RM) with minimum yield loss (scale:0) and the susceptible mutant (SM) with maximum loss (scale:4) were used for metabolomic analysis through GC-MS. PM infection induced expression of 66 metabolites representing 32 functional classes. The number of unique classes in RM and SM was 13 and 11, respectively, while eight were common. A fold change in the common metabolites indicated an enhanced accumulation of amine, alcohol, and ester in RM. Along with pathogen-induced defensive metabolites, RM produced silane and fluorene, whose biological significance in disease resistance is unknown. Though SM expressed defence-related bio-molecules, it failed to yield better.
Journal of Plant Physiology, 2014
Artemisinin, an antimalarial endoperoxide sesquiterpene, is synthesized in glandular trichomes of... more Artemisinin, an antimalarial endoperoxide sesquiterpene, is synthesized in glandular trichomes of Artemisia annua L. A number of other enzymes of terpene metabolism utilize intermediates of artemisinin biosynthesis, such as isopentenyl and farnesyl diphosphate, and may thereby influence the yield of artemisinin. In order to study the expression of such enzymes, we have cloned the promoter regions of some enzymes and fused them to -glucuronidase (GUS). In this study, we have investigated the expression of the monoterpene synthase linalool synthase (LIS) using transgenic A. annua carrying the GUS gene under the control of the LIS promoter. The 652 bp promoter region was cloned by the genome walker method. A number of putative cis-acting elements were predicted indicating that the LIS is driven by a complex regulation mechanism. Transgenic plants carrying the promoter-GUS fusion showed specific expression of GUS in T-shaped trichomes (TSTs) but not in glandular secretory trichomes, which is the site for artemisinin biosynthesis. GUS expression was observed at late stage of flower development in styles of florets and in TSTs and guard cells of basal bracts. GUS expression after wounding showed that LIS is involved in plant responsiveness to wounding. Furthermore, the LIS promoter responded to methyl jasmonate (MeJA). These results indicate that the promoter carries a number of cis-acting regulatory elements involved in the tissue-specific expression of LIS and in the response of the plant to wounding and MeJA treatment. Southern blot analysis indicated that the GUS gene was integrated in the A. annua genome as single or multi copies in different transgenic lines. Promoter activity analysis by qPCR showed that both the wild-type and the recombinant promoter are active in the aerial parts of the plant while only the recombinant promoter was active in roots. Due to the expression in TSTs but not in glandular trichomes, it may be concluded that LIS expression will most likely have little or no effect on artemisinin production.
PLOS ONE, Nov 22, 2013
In order to better understand the influence of sesquiterpene synthases on artemisinin yield in Ar... more In order to better understand the influence of sesquiterpene synthases on artemisinin yield in Artemisia annua, the expression of some sesquiterpene synthases has been studied using transgenic plants expressing promoter-GUS fusions. The cloned promoter sequences were 923, 1182 and 1510 bp for b-caryophyllene (CPS), epi-cedrol (ECS) and b-farnesene (FS) synthase, respectively. Prediction of cis-acting regulatory elements showed that the promoters are involved in complex regulation of expression. Transgenic A. annua plants carrying promoter-GUS fusions were studied to elucidate the expression pattern of the three sesquiterpene synthases and compared to the previously studied promoter of amorpha-4,11-diene synthase (ADS), a key enzyme of artemisinin biosynthesis. The CPS and ECS promoters were active in T-shaped trichomes of leaves and stems, basal bracts of flower buds and also in some florets cells but not in glandular secretory trichome while FS promoter activity was only observed in leaf cells and trichomes of transgenic shoots. ADS, CPS, ECS and FS transcripts were induced by wounding in a time depended manner. The four sesquiterpene synthases may be involved in responsiveness of A. annua to herbivory. Methyl jasmonate treatment triggered activation of the promoters of all four sesquiterpene synthases in a time depended manner. Southern blot result showed that the GUS gene was inserted into genomic DNA of transgenic lines as a single copy or two copies. The relative amounts of CPS and ECS as well as germacrene A synthase (GAS) transcripts are much lower than that of ADS transcript. Consequently, down-regulation of the expression of the CPS, ECS or GAS gene may not improve artemsinin yield. However, blocking the expression of FS may have effects on artemisinin production.
Conservation Genetics, Nov 28, 2008
Abstract We developed and characterized 8 polymorphic microsatellite loci or simple sequence repe... more Abstract We developed and characterized 8 polymorphic microsatellite loci or simple sequence repeats from the expressed sequence tags database of the medicinal fungus, Antrodia cinnamomea. Variability was assessed in a panel of 23 strains collected from ...
Plant Cell Reports, May 8, 2012
Artemisia annua L. produces a number of sesquiterpene synthases, which catalyze the conversion of... more Artemisia annua L. produces a number of sesquiterpene synthases, which catalyze the conversion of farnesyl diphosphate to various sesquiterpenes. The cDNAs encoding amorpha-4,11-diene synthase (ADS), a key enzyme in the artemisinin biosynthesis, and epi-cedrol synthase (ECS), a complex sesquiterpene cyclization synthase, were cloned into Cowpea mosaic virus-based viral vector (pEAQ-HT) with Kozak consensus motif and C-terminal histidine tag. The plasmids were transformed into Agrobacterium LBA4404 and, agroinfiltrated into Nicotiana benthamiana leaves along with vector (pJL3:p19) containing Tomato bushy stunt virus post-transcriptional gene silencing suppressor. Quantitative PCR was carried out to measure the transcript levels at 0, 3, 6, 9, 12 and 15 days post-infiltration (dpi). The highest relative expression was observed at 9 dpi for both genes. Transiently expressed recombinant proteins of ADS and ECS were confirmed by SDS-PAGE and western blot. Recombinant proteins were extracted from 9 dpi leaves and purified by immobilized metal ion affinity chromatography using histidine tag, which produced yields of 90 and 96 mg kg⁻¹ fresh weight of leaves for ADS and ECS, respectively. Activities of the purified enzymes were assayed using gas chromatography-mass spectrometry for product identification and quantification using valencene as internal standard. The recombinant ADS and ECS converted farnesyl diphosphate into amorpha-4,11-diene (97 %) and epi-cedrol (96 %) as the major products, respectively. The purified enzymes exhibited the specific activity of 0.002 and 0.01 μmol min⁻¹ mg⁻¹ protein for ADS and ECS, respectively. The apparent k(cat) values were 2.1 × 10⁻³ s⁻¹ and 11 × 10⁻³ s⁻¹ for ADS and ECS, respectively. Agroinfiltration of leaves of Nicotiana bentamiana can be used to produce recombinant biosynthetic enzymes as exemplified by two sesquiterpene synthases from Artemisia annua in relatively high yields.
Frontiers in Plant Science, Jan 18, 2023
The wild species field cress (Lepidium campestre) has the potential to become a novel cover and o... more The wild species field cress (Lepidium campestre) has the potential to become a novel cover and oilseed crop for the Nordic climate. Its seed oil is however currently unsuitable for most food, feed, and industrial applications, due to the high contents of polyunsaturated fatty acids (PUFAs) and erucic acid (C22:1). As the biosynthesis of these undesirable fatty acids is controlled by a few well-known major dominant genes, knockout of these genes using CRISPR/Cas9 would thus be more effective in improving the seed oil quality. In order to increase the level of the desirable oleic acid (C18:1), and reduce the contents of PUFAs and C22:1, we targeted three important genes FATTY ACID ELONGASE1 (FAE1), FATTY ACID DESATURASE2 (FAD2), and REDUCED OLEATE DESATURASE1 (ROD1) using a protoplast-based CRISPR/Cas9 gene knockout system. By knocking out FAE1, we obtained a mutated line with almost no C22:1, but an increase in C18:1 to 30% compared with 13% in the wild type. Knocking out ROD1 resulted in an increase of C18:1 to 23%, and a moderate, but significant, reduction of PUFAs. Knockout of FAD2, in combination with heterozygous FAE1fae1 genotype, resulted in mutated lines with up to 66% C18:1, very low contents of PUFAs, and a significant reduction of C22:1. Our results clearly show the potential of CRISPR/Cas9 for rapid trait improvement of field cress which would speed up its domestication process. The mutated lines produced in this study can be used for further breeding to develop field cress into a viable crop.
Plants
Mung bean, a legume, is sensitive to abiotic stresses at different growth stages, and its yield p... more Mung bean, a legume, is sensitive to abiotic stresses at different growth stages, and its yield potential is affected by drought and high-temperature stress at the sensitive stage. Melatonin is a multifunctional hormone that plays a vital role in plant stress defense mechanisms. This study aimed to evaluate the efficiency of melatonin under individual and combined drought and high-temperature stress in mung bean. An experiment was laid out with five treatments, including an exogenous application of 100 µM melatonin as a seed treatment, foliar spray, and a combination of both seed treatment and foliar spray, as well as absolute control (ambient condition) and control (stress without melatonin treatment). Stresses were imposed during the mung bean’s reproductive stage (31–40 DAS) for ten days. Results revealed that drought and high-temperature stress significantly decreased chlorophyll index, Fv/Fm ratio, photosynthetic rate, stomatal conductance, and transpiration rate through increa...
Metabolites, Jan 23, 2023
Horse gram (Macrotyloma uniflorum (Lam.) Verdc.) is an under-utilized legume grown in India. It i... more Horse gram (Macrotyloma uniflorum (Lam.) Verdc.) is an under-utilized legume grown in India. It is a good source of protein, carbohydrates, dietary fiber, minerals, and vitamins. We screened 252 horse gram germplasm accessions for horse gram yellow mosaic virus resistance using the percent disease index and scaling techniques. The percentage values of highly resistant, moderately resistant, moderately susceptible, susceptible, and highly susceptible were 0.34, 13.89, 38.89, 46.43, and 0.34, respectively. Repetitive trials confirmed the host-plant resistance levels, and yield loss was assessed. The present disease index ranged from 1.2 to 72.0 and 1.2 to 73.0 during the kharif and rabi seasons of 2018, respectively. The maximum percent yield loss was noticed in the HS (75.0 -89.4), while HR possessed the minimum (1.2-2.0). The methanolic leaf extracts of highly resistant and highly susceptible genotypes with essential controls were subjected to gas chromatography-mass spectrometry analysis. Differential accumulation of metabolites was noticed, and a total of 81 metabolites representing 26 functional groups were identified. Both highly resistant and susceptible genotypes harbored eight unique classes, while ten biomolecules were common. The hierarchical cluster analysis indicated a distinct metabolite profile. Fold change in the common metabolites revealed an enhanced accumulation of sugars, alkanes, and carboxylic acids in the highly resistant genotype. The principal component analysis plots explained 93.7% of the variation. The metabolite profile showed a significant accumulation of three anti-viral (octadecanoic acid, diphenyl sulfone, and 2-Aminooxazole), one insecticidal (9,10-Secocholesta-5,7,10(19)-triene-3,24,25-triol), one antifeedant (cucurbitacin B), and six metabolites with unknown biological function in the highly resistant genotype.
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Papers by Selvaraju Kanagarajan