@article {7318, title = {Techno-commercial application of microencapsulated basil oil and thyme oil and their antibacterial activity against aquatic pathogens [EM Facility]}, journal = {International Journal of Fisheries and Aquatic Studies 2022}, volume = {10(5)}, year = {2022}, month = {23/06/2022}, pages = {101-106}, type = {Journal Article}, chapter = {101}, abstract = {

A complete study was conducted through manufacturing of Microencapsulation of Basil Oil Powder (MEBOP) and Microencapsulation of Thyme Oil Powder (METOP) to evaluate the anti-bacterial activity against aquatic pathogens. An oil-in-water emulsion were prepared using gum with dextrin and the resultant matrix was spray dried with the average yield of 45\% w/w of MEBOP and 50\% w/w of METOP. Total oil content in the encapsulated powder in both were found to be 24.39\% w/w of MEBOP 25.80\% w/w of METOP respectively. Scanning Electron Microscopic (SEM) analysis was also done to confirm the encapsulation of oil and study possible structure of matrix. Microencapsulated powders were further subjected for antibacterial activity against aquaculture pathogens like Vibrio harvey, Vibrio parahaemolyticus, Bacillus cereus, Aeromonas sp, E. coli and Salmonella sp. Results were compared against the respective oils at different concentrations. Phyto- constituents from Basil oil and Thyme oil like Thymol, Linalool, and Methyl chavicol were quantified using HPLC (High-Performance Liquid Chromatography) and GC (Gas Chromatography) which may be responsible for this activity. Furthermore, stress studies were conducted at 60 {\textdegree}C to understand the stability of Encapsulated oil powders to this and its commercial usage in formulations.

}, keywords = {anti-bacterial, aqua gut health, Microencapsulated oil powder, stress study}, issn = {E-ISSN: 2347-5129}, doi = {https://doi.org/10.22271/fish.2022.v10.i5b.2730}, url = {https://www.fisheriesjournal.com/archives/2022/vol10issue5/PartB/10-4-44-468.pdf}, author = {Sampath, M and Muguli, Ganesh and Pai, Sameer and Babu, UV} } @article {6781, title = {Techno-commercial application of microencapsulated basil oil and thyme oil and their antibacterial activity against aquatic pathogens [Electron Microscopy Facility]}, journal = {International Journal of Fisheries and Aquatic Studies}, volume = {10}, year = {2022}, month = {10/2022}, chapter = {101-106}, abstract = {

A complete study was conducted through manufacturing of Microencapsulation of Basil Oil Powder (MEBOP) and Microencapsulation of Thyme Oil Powder (METOP) to evaluate the anti-bacterial activity against aquatic pathogens. An oil-in-water emulsion were prepared using gum with dextrin and the resultant matrix was spray dried with the average yield of 45\% w/w of MEBOP and 50\% w/w of METOP. Total oil content in the encapsulated powder in both were found to be 24.39\% w/w of MEBOP 25.80\% w/w of METOP respectively. Scanning Electron Microscopic (SEM) analysis was also done to confirm the encapsulation of oil and study possible structure of matrix. Microencapsulated powders were further subjected for antibacterial activity against aquaculture pathogens like Vibrio harvey, Vibrio parahaemolyticus, Bacillus cereus, Aeromonas sp, E. coli and Salmonella sp. Results were compared against the respective oils at different concentrations. Phyto- constituents from Basil oil and Thyme oil like Thymol, Linalool, and Methyl chavicol were quantified using HPLC (High-Performance Liquid Chromatography) and GC (Gas Chromatography) which may be responsible for this activity. Furthermore, stress studies were conducted at 60 {\textdegree}C to understand the stability of Encapsulated oil powders to this and its commercial usage in formulations.

}, keywords = {anti-bacterial, aqua gut health, Microencapsulated oil powder, stress study}, issn = {2394-0506}, doi = {10.22271/fish.2022.v10.i5b.2730}, url = {https://www.fisheriesjournal.com/archives/2022/vol10issue5/PartB/10-4-44-468.pdf}, author = {Sampath M and Ganesh Muguli and Sameer Pai and Babu UV} } @article {2733, title = {Transcriptome analysis at mid-stage seed development in litchi with contrasting seed size [Next Gen Genomics Facility]}, journal = {3 Biotech }, volume = {12}, year = {2022}, month = {01/2022}, abstract = {

Litchi is a sub-tropical fruit crop with genotypes that bear fruits with variable seed size. Small seed size is a desirable trait\ in litchi, as it improves consumers{\textquoteright} preference and facilitates fruit processing. Seed specific transcriptome analysis was performed in two litchi genotypes with contrasting seed size to identify the genes associated with seed development. The transcriptomic sequence data from seeds at mid-development stages (16{\textendash}28\ days after anthesis) were de-novo assembled into 1,39,608 Trinity transcripts. Out of these, 6325 transcripts expressed differentially between the two contrasting genotypes. Several putative genes for salicylic acid, jasmonic acid and brassinosteriod pathways were down-regulated in seeds of the small-seeded litchi. The putative regulators of seed maturation and seed storage were down-regulated in the small-seeded genotype. Embryogenesis, cell\ expansion, seed size and stress related Trinity transcripts exhibited differential expression. Further studies on gene characterization will reveal the early regulators of seed size in litchi.

}, keywords = {Embryogenesis, Litchi, Seed size, Small seed, Transcriptome}, doi = {doi.org/10.1007/s13205-021-03098-8}, url = {https://link.springer.com/article/10.1007/s13205-021-03098-8$\#$citeas}, author = {Ashish K. Pathak and Sudhir P. Singh and Ritika Sharma and Vishal Nath and Rakesh Tuli} } @article {1639, title = {Transplantation of retinal pigment epithelium and photoreceptors generated concomitantly via small molecule-mediated differentiation rescues visual function in rodent models of retinal degeneration [Eyestem Research Pvt. Ltd., a C-CAMP Startup]}, journal = {Stem Cell Res Ther}, volume = {12}, year = {2021}, month = {2021 Jan 19}, pages = {70}, abstract = {

BACKGROUND: Age-related macular degeneration (AMD) is a result of degeneration/damage of the retinal pigment epithelium (RPE) while retinitis pigmentosa (RP), an inherited early-onset disease, results from premature loss of photoreceptors. A promising therapeutic approach for both is the replacement of lost/damaged cells with human induced pluripotent stem cell (hiPSC)-derived retinal cells.

METHODS: The aim of this study was to investigate the in vivo functionality of RPE and photoreceptor progenitor (PRP) cells derived from a clinical-grade hiPSC line through a unified protocol. De novo-generated RPE and PRP were characterized extensively to validate their identity, purity, and potency.

RESULTS: RPE expressed tight junction proteins, showed pigmentation and ciliation, and secreted polarization-related factors vascular endothelial growth factor (VEGF) and pigment epithelium-derived factor (PEDF). PRP expressed neural retina proteins and cone and rod markers, and responded to KCl-induced polarization. Transcriptomic analysis demonstrated an increase in the expression of mature retinal tissue-specific genes coupled with concomitant downregulation of genes from undesired lineages. RPE transplantation rescued visual function in RCS rats shown via optokinetic tracking and photoreceptor rescue. PRP transplantation improved light perception in NOD.SCID-rd1 mice, and positive electroretinography signals indicated functional photoreceptor activity in the host{\textquoteright}s outer nuclear layer. Graft survival and integration were confirmed using immunohistochemistry, and no animals showed teratoma formation or any kind of ectopic growth in the eye.

CONCLUSIONS: To our knowledge, this is the first demonstration of a unified, scalable, and GMP-adaptable protocol indicating strong animal efficacy and safety data with hiPSC-derived RPE and PRP cells. These findings provide robust proof-of-principle results for IND-enabling studies to test these potential regenerative cell therapies in patients.

}, issn = {1757-6512}, doi = {10.1186/s13287-021-02134-x}, author = {Surendran, Harshini and Nandakumar, Swapna and Reddy K, Vijay Bhaskar and Stoddard, Jonathan and Mohan K, Varsha and Upadhyay, Pramod K and McGill, Trevor J and Pal, Rajarshi} } @article {1677, title = {The truth about scats and dogs: Next-generation sequencing and spatial capture{\textendash}recapture models offer opportunities for conservation monitoring of an endangered social canid [Next Gen Genomics Facility (INT)].}, journal = {Biological Conservation}, volume = {256}, year = {2021}, pages = {109028}, abstract = {

Obtaining accurate population counts of endangered species is central to conservation biology, with implications for gaining ecological insights, informing management strategies, and judicial use of conservation funds. Despite decades of progress in methodological developments in the realm of population ecology, reliable density estimates are unavailable for many species of conservation concern. The dhole (Asiatic wild dog Cuon alpinus) is one such endangered large carnivore found in the tropical forests of south and southeast Asia. Here, we (i) develop next-generation sequencing resources to identify individual dholes from genetic samples, (ii) apply these methods to identify individuals in the wild, from scat (fecal) samples collected through systematic field surveys and (iii) generate reliable estimates of dhole densities in Wayanad Wildlife Sanctuary (Western Ghats, India) using Spatial Capture{\textendash}Recapture {\textquoteleft}SCR{\textquoteright} models. We estimate dhole densities to be 12{\textendash}14.2 individuals/100 sq. km based on a set of SCR models, with \ 50 individuals within Wayanad{\textquoteright}s administrative boundary. Our study presents a methodological improvement in generating population estimates of an important apex predator while also offering ecologically informative insights on a species in dire need of science-based management efforts. Replicating this study across connected reserves and over time can serve as a unified framework for understanding population dynamics, population structures, landscape connectivity and metapopulation-level conservation requirements. We propose that the approach presented here may be adopted as an economically and logistically feasible protocol for conservation monitoring of dholes and other ecologically important species plagued by similar issues of data-deficiency, and insufficient funding and resources.

}, keywords = {Carnivores, Conservation monitoring, Genetic markers, Non-invasive surveys, Population estimation, Single nucleotide polymorphisms, Tropics}, issn = {0006-3207}, doi = {https://doi.org/10.1016/j.biocon.2021.109028}, url = {https://www.sciencedirect.com/science/article/pii/S000632072100080X}, author = {Arjun Srivathsa and Ryan G. Rodrigues and Kok Ben Toh and Arun Zachariah and Ryan W. Taylor and Madan K. Oli and Uma Ramakrishnan} } @article {1343, title = {Tracing the origin of olive ridley turtles entangled in ghost nets in the Maldives: A phylogeographic assessment of populations at risk [Next Gen Genomics Facility]}, journal = {Biological Conservation}, volume = {245}, year = {2020}, pages = {108499}, abstract = {

Abandoned, lost or discarded fishing nets, (ghost nets) represent a major threat to marine vertebrates. However, thorough assessments of their impact on threatened species are largely missing. In the Maldives, olive ridley sea turtles (Lepidochelys olivacea) are frequently caught in ghost nets however the archipelago does not support a significant nesting population. Our aim in this study was to determine the origin of olive ridleys entangled in ghost nets found in the Maldives and evaluate potential impacts on respective source populations. Based on a citizen science and conservation program, we recorded 132 olive ridley turtles entangled in ghost nets in just one year. Genetic analyses (mtDNA) of entangled individuals and of potential source populations revealed that most captured olive ridleys originated from Sri Lanka and eastern India. Oman could be excluded as source population, even during the prevalence of the south west monsoon. Based on our results and already available published literature, we were able to estimate that the recorded ghost net entanglements accounted for a relatively small amount (0.48\%) of the eastern Indian population. However, the entangled turtles accounted for a much larger percentage (41\%) of the Sri Lankan populations. However, it should be noted that our estimates of population-level mortality are linked to substantial uncertainty due to the lack of reliable information on population dynamics. Consequently, any precautionary protection measures applied should be complemented with improved quantification of turtle recruitment and life-stage specific mortalities.

}, keywords = {Citizen science, Entanglement, Ghost nets, mtDNA, Phylogenetics, Plastics}, issn = {0006-3207}, doi = {https://doi.org/10.1016/j.biocon.2020.108499}, url = {http://www.sciencedirect.com/science/article/pii/S0006320719313874}, author = {Martin Stelfox and Alfred Burian and Kartik Shanker and Alan F. Rees and Claire Jean and Ma{\"\i}a S. Willson and Nashwa Ahmed Manik and Michael Sweet} } @article {1524, title = {Tracking the time-dependent and tissue-specific processes of arsenic accumulation and stress responses in rice (Oryza sativa L.) [Mass Spectrometry - Metabolomics Facility]}, journal = {Journal of Hazardous Materials}, year = {2020}, pages = {124307}, abstract = {

The present study analysed time (0.5h to 24h) and tissue [roots, old leaves (OL) and young leaves (YL)] dependent nature of arsenic (As) accumulation and ensuing responses in two contrasting varieties of rice (Oryza sativa L.); Pooja (tolerant) and CO-50 (moderately sensitive). Arsenic accumulation was 5.4-, 4.7- and 7.3-fold higher at 24h in roots, OL and YL, respectively of var. CO-50 than that in var. Pooja. Arsenic accumulation in YL depicted a delayed accumulation; at 2h onwards in var. Pooja (0.23{\textmu}gg-1 dw) while at 1h onwards in var. CO50 (0.26{\textmu}gg-1 dw). The responses of oxidative stress parameters, antioxidant enzymes, metabolites and ions were also found to be tissue- and time-dependent and depicted differential pattern in the two varieties. Among hormone, salicylic acid and abscisic acid showed variable response in var. Pooja and var. CO-50. Metabolite analysis depicted an involvement of various metabolites in As stress responses of two varieties. In conclusion, an early sensing of the As stress, proper coordination of hormones, biochemical responses, ionic and metabolic profiles allowed var. Pooja to resist As stress and reduce As accumulation more effectively as compared to that of var. CO-50.

}, keywords = {Arsenic, Hormones, Metabolites, ROS, Signalling, Stress}, issn = {0304-3894}, doi = {https://doi.org/10.1016/j.jhazmat.2020.124307}, url = {http://www.sciencedirect.com/science/article/pii/S0304389420322974}, author = {Poonam Yadav and Sudhakar Srivastva and Tanmayi Patil and Rishiraj Raghuvanshi and Ashish K. Srivastava and Penna Suprasanna} } @article {1228, title = {Transcriptome analysis reveals plasticity in gene regulation due to environmental cues in Primula sikkimensis, a high altitude plant species [Next Gen Genomics Facility (INT)].}, journal = {BMC Genomics}, volume = {20}, year = {2019}, month = {2019 Dec 17}, pages = {989}, abstract = {

BACKGROUND: Studying plasticity in gene expression in natural systems is crucial, for predicting and managing the effects of climate change on plant species. To understand the contribution of gene expression level variations to abiotic stress compensation in a Himalaya plant (Primula sikkimensis), we carried out a transplant experiment within (Ambient), and beyond (Below Ambient and Above Ambient) the altitudinal range limit of species. We sequenced nine transcriptomes (three each from each altitudinal range condition) using Illumina sequencing technology. We compared the fitness variation of transplants among three transplant conditions.

RESULTS: A large number of significantly differentially expressed genes (DEGs) between below ambient versus ambient (109) and above ambient versus ambient (85) were identified. Transcripts involved in plant growth and development were mostly up-regulated in below ambient conditions. Transcripts involved in signalling, defence, and membrane transport were mostly up-regulated in above ambient condition. Pathway analysis revealed that most of the genes involved in metabolic processes, secondary metabolism, and flavonoid biosynthesis were differentially expressed in below ambient conditions, whereas most of the genes involved in photosynthesis and plant hormone signalling were differentially expressed in above ambient conditions. In addition, we observed higher reproductive fitness in transplant individuals at below ambient condition compared to above ambient conditions; contrary to what we expect from the cold adaptive P. sikkimensis plants.

CONCLUSIONS: We reveal P. sikkimensis{\textquoteright}s capacity for rapid adaptation to climate change through transcriptome variation, which may facilitate the phenotypic plasticity observed in morphological and life history traits. The genes and pathways identified provide a genetic resource for understanding the temperature stress (both the hot and cold stress) tolerance mechanism of P. sikkimensis in their natural environment.

}, issn = {1471-2164}, doi = {10.1186/s12864-019-6354-1}, author = {Gurung, Priya Darshini and Upadhyay, Atul Kumar and Bhardwaj, Pardeep Kumar and Sowdhamini, Ramanathan and Ramakrishnan, Uma} } @article {1084, title = {The transcriptome enables the identification of candidate genes behind medicinal value of Drumstick tree (Moringa oleifera) [Next Gen Genomics Facility (INT)].}, journal = {Genomics}, year = {2019}, month = {2019 Apr 29}, abstract = {

Moringa oleifera is a plant well-known for its nutrition value, drought resistance and medicinal properties. cDNA libraries from five different tissues (leaf, root, stem, seed and flower) of M. oleifera cultivar Bhagya were generated and sequenced. We developed a bioinformatics pipeline to assemble transcriptome, along with the previously published M. oleifera genome, to predict 17,148 gene models. Few candidate genes related to biosynthesis of secondary metabolites, vitamins and ion transporters were identified. Expressions were further confirmed by real-time quantitative PCR experiments for few promising leads. Quantitative estimation of metabolites, as well as elemental analysis, was also carried out to support our observations. Enzymes in the biosynthesis of vitamins and metabolites like quercetin and kaempferol are highly expressed in leaves, flowers and seeds. The expression of iron transporters and calcium storage proteins were observed in root and leaves. In general, leaves retain the highest amount of small molecules of interest.

}, issn = {1089-8646}, doi = {10.1016/j.ygeno.2019.04.014}, author = {Pasha, Shaik Naseer and Shafi, K Mohamed and Joshi, Adwait G and Meenakshi, Iyer and Harini, K and Mahita, Jarjapu and Sajeevan, Radha Sivarajan and Karpe, Snehal D and Ghosh, Pritha and Nitish, Sathyanarayanan and Gandhimathi, A and Mathew, Oommen K and Prasanna, Subramanian Hari and Malini, Manoharan and Mutt, Eshita and Naika, Mahantesha and Ravooru, Nithin and Rao, Rajas M and Shingate, Prashant N and Sukhwal, Anshul and Sunitha, Margaret S and Upadhyay, Atul K and Vinekar, Rithvik S and Sowdhamini, Ramanathan} } @article {1020, title = {Transcriptomics analysis of propiconazole-treated Cochliobolus sativus reveals new putative azole targets in the plant pathogen [Next Gen Genomics Facility].}, journal = {Funct Integr Genomics}, year = {2019}, month = {2019 Mar 06}, abstract = {

Cochliobolus sativus (anamorph: Bipolaris sorokiniana) is a filamentous fungus from the class Dothideomycetes. It is a pathogen of cereals including wheat and barley, and causes foliar spot blotch, root rot, black point on grains, head blight, leaf blight, and seedling blight diseases. Annual yields of these economically important cereals are severely reduced due to this pathogen attack. Evolution of fungicide resistant pathogen strains, availability of a limited number of potent antifungal compounds, and their efficacy are the acute issues in field management of phytopathogenic fungi. Propiconazole is a widely used azole fungicide to control the disease in fields. The known targets of azoles are the demethylase enzymes involved in ergosterol biosynthesis. Nonetheless, azoles have multiple modes of action, some of which have not been explored yet. Identifying the off-target effects of fungicides by dissecting gene expression profiles in response to them can provide insights into their modes of action and possible mechanisms of fungicide resistance. Moreover it can also reveal additional targets for development of new fungicides. Hence, we analyzed the global gene expression profile of C. sativus on exposure to sub-lethal doses of propiconazole in a time series. The gene expression patterns were confirmed using quantitative reverse transcriptase PCR (qRT-PCR). This study revealed overexpression of target genes from the sterol biosynthesis pathway supporting the reported mode of resistance against azoles. In addition, some new potential targets have also been identified, which could be explored to develop new fungicides and plant protection strategies.

}, issn = {1438-7948}, doi = {10.1007/s10142-019-00660-9}, author = {Somani, Deepika and Adhav, Ragini and Prashant, Ramya and Kadoo, Narendra Y} } @article {532, title = {Targeted, Site-specific quantitation of N- and O-glycopeptides using (18)O-labeling and product ion based mass spectrometry.}, journal = {Glycoconj J}, volume = {34}, year = {2017}, month = {2017 Feb}, pages = {95-105}, abstract = {

The site-specific quantitation of N- and O-glycosylation is vital to understanding the function(s) of different glycans expressed at a given site of a protein under physiological and disease conditions. Most commonly used precursor ion intensity based quantification method is less accurate and other labeled methods are expensive and require enrichment of glycopeptides. Here, we used glycopeptide product (y and Y0) ions and (18)O-labeling of C-terminal carboxyl group as a strategy to obtain quantitative information about fold-change and relative abundance of most of the glycoforms attached to the glycopeptides. As a proof of concept, the accuracy and robustness of this targeted, relative quantification LC-MS method was demonstrated using Rituximab. Furthermore, the N-glycopeptide quantification results were compared with a biosimilar of Rituximab and validated with quantitative data obtained from 2-AB-UHPLC-FL method. We further demonstrated the intensity fold-change and relative abundance of 46 unique N- and O-glycopeptides and aglycopeptides from innovator and biosimilar samples of Etanercept using both the normal-MS and product ion based quantitation. The results showed a very similar site-specific expression of N- and O-glycopeptides between the samples but with subtle differences. Interestingly, we have also been able to quantify macro-heterogeneity of all N- and O-glycopetides of Etanercept. In addition to applications in biotherapeutics, the developed method can also be used for site-specific quantitation of N- and O-glycopeptides and aglycopeptides of glycoproteins with known glycosylation pattern.

}, issn = {1573-4986}, doi = {10.1007/s10719-016-9733-8}, author = {Srikanth, Jandhyam and Agalyadevi, Rathinasamy and Babu, Ponnusamy} } @article {8889, title = {Transcriptional transitions in Alphonso mango (Mangifera indica L.) during fruit development and ripening explain its distinct aroma and shelf life characteristics [Next Gen Genomics Facility]}, journal = {Sci Rep}, volume = {7}, year = {2017}, month = {2017 Aug 18}, pages = {8711}, abstract = {

Alphonso is known as the "King of mangos" due to its unique flavor, attractive color, low fiber pulp and long shelf life. We analyzed the transcriptome of Alphonso mango through Illumina sequencing from seven stages of fruit development and ripening as well as flower. Total transcriptome data from these stages ranged between 65 and 143 Mb. Importantly, 20,755 unique transcripts were annotated and 4,611 were assigned enzyme commission numbers, which encoded 142 biological pathways. These included ethylene and flavor related secondary metabolite biosynthesis pathways, as well as those involved in metabolism of starch, sucrose, amino acids and fatty acids. Differential regulation (p-value <= 0.05) of thousands of transcripts was evident in various stages of fruit development and ripening. Novel transcripts for biosynthesis of mono-terpenes, sesqui-terpenes, di-terpenes, lactones and furanones involved in flavor formation were identified. Large number of transcripts encoding cell wall modifying enzymes was found to be steady in their expression, while few were differentially regulated through these stages. Novel 79 transcripts of inhibitors of cell wall modifying enzymes were simultaneously detected throughout Alphonso fruit development and ripening, suggesting controlled activity of these enzymes involved in fruit softening.

}, keywords = {Cell Wall, Enzyme Inhibitors, Flowers, Fruit, Gene Expression Profiling, Gene Expression Regulation, Developmental, Gene Expression Regulation, Plant, Gene Ontology, Genes, Plant, Glycoside Hydrolases, Mangifera, Odorants, Phylogeny, Reproducibility of Results, RNA, Messenger, Transcription, Genetic, Transcriptome}, issn = {2045-2322}, doi = {10.1038/s41598-017-08499-5}, author = {Deshpande, Ashish B and Anamika, Krishanpal and Jha, Vineet and Chidley, Hemangi G and Oak, Pranjali S and Kadoo, Narendra Y and Pujari, Keshav H and Giri, Ashok P and Gupta, Vidya S} } @article {688, title = {Transcriptome profiling of Elettaria cardamomum (L.) Maton (small cardamom).}, journal = {Genom Data}, volume = {11}, year = {2017}, month = {2017 Mar}, pages = {102-103}, abstract = {

Elettaria cardamomum (L.) Maton, known as {\textquoteright}queen of spices, is a perennial herbaceous monocot of the family Zingiberaceae, native to southern India. Cardamom is an economically valuable spice crop and used widely in culinary and medicinal purposes. In the present study, using Ion Proton RNA sequencing technology, we performed transcriptome sequencing and de novo transcriptome assembly of a wild and five cultivar genotypes of cardamom. RNA-seq generated a total of 22,811,983 (92 base) and 24,889,197 (75 base) raw reads accounting for approximately 8.21GB and 7.65GB of sequence data for wild and cultivar genotypes of cardamom respectively. The raw data were submitted to SRA database of NCBI under the accession numbers SRX1141272 (wild) and SRX1141276 (cultivars). The raw reads were quality filtered and assembled using MIRA assembler resulted with 112,208 and 264,161contigs having N50 value 616 and 664 for wild and cultivar cardamom respectively. The assembled unigenes were functionally annotated using several databases including PlantCyc for pathway annotation. This work represents the first report on cardamom transcriptome sequencing. In order to generate a comprehensive reference transcriptome, we further assembled the raw reads of wild and cultivar genotypes which might enrich the plant transcriptome database and trigger advanced research in cardamom genomics.

}, issn = {2213-5960}, doi = {10.1016/j.gdata.2016.12.013}, author = {Nadiya, F and Anjali, N and Thomas, Jinu and Gangaprasad, A and Sabu, K K} } @article {8890, title = {Transcriptomic resources for the medicinal legume Mucuna pruriens: de novo transcriptome assembly, annotation, identification and validation of EST-SSR markers [Next Gen Genomics Facility]}, journal = {BMC Genomics}, volume = {18}, year = {2017}, month = {2017 May 25}, pages = {409}, abstract = {

BACKGROUND: The medicinal legume Mucuna pruriens (L.) DC. has attracted attention worldwide as a source of the anti-Parkinson{\textquoteright}s drug L-Dopa. It is also a popular green manure cover crop that offers many agronomic benefits including high protein content, nitrogen fixation and soil nutrients. The plant currently lacks genomic resources and there is limited knowledge on gene expression, metabolic pathways, and genetics of secondary metabolite production. Here, we present transcriptomic resources for M. pruriens, including a de novo transcriptome assembly and annotation, as well as differential transcript expression analyses between root, leaf, and pod tissues. We also develop microsatellite markers and analyze genetic diversity and population structure within a set of Indian germplasm accessions.

RESULTS: One-hundred ninety-one million two hundred thirty-three thousand two hundred forty-two\ bp cleaned reads were assembled into 67,561 transcripts with mean length of 626\ bp and N50 of 987\ bp. Assembled sequences were annotated using BLASTX against public databases with over 80\% of transcripts annotated. We identified 7,493 simple sequence repeat (SSR) motifs, including 787 polymorphic repeats between the parents of a mapping population. 134 SSRs from expressed sequenced tags (ESTs) were screened against 23 M. pruriens accessions from India, with 52 EST-SSRs retained after quality control. Population structure analysis using a Bayesian framework implemented in fastSTRUCTURE showed nearly similar groupings as with distance-based (neighbor-joining) and principal component analyses, with most of the accessions clustering per geographical origins. Pair-wise comparison of transcript expression in leaves, roots and pods identified 4,387 differentially expressed transcripts with the highest number occurring between roots and leaves. Differentially expressed transcripts were enriched with transcription factors and transcripts annotated as belonging to secondary metabolite pathways.

CONCLUSIONS: The M. pruriens transcriptomic resources generated in this study provide foundational resources for gene discovery and development of molecular markers. Polymorphic SSRs identified can be used for genetic diversity, marker-trait analyses, and development of functional markers for crop improvement. The results of differential expression studies can be used to investigate genes involved in L-Dopa synthesis and other key metabolic pathways in M. pruriens.

}, keywords = {Data Mining, Expressed Sequence Tags, Gene Expression Profiling, Microsatellite Repeats, Molecular Sequence Annotation, Mucuna, Polymorphism, Genetic, RNA, Messenger, Transcription Factors}, issn = {1471-2164}, doi = {10.1186/s12864-017-3780-9}, author = {Sathyanarayana, N and Pittala, Ranjith Kumar and Tripathi, Pankaj Kumar and Chopra, Ratan and Singh, Heikham Russiachand and Belamkar, Vikas and Bhardwaj, Pardeep Kumar and Doyle, Jeff J and Egan, Ashley N} } @article {507, title = {Tailor-made ezrin actin binding domain to probe its interaction with actin in-vitro. [Protein Technology Core]}, journal = {PLoS One}, volume = {10}, year = {2015}, month = {2015}, pages = {e0123428}, abstract = {

Ezrin, a member of the ERM (Ezrin/Radixin/Moesin) protein family, is an Actin-plasma membrane linker protein mediating cellular integrity and function. In-vivo study of such interactions is a complex task due to the presence of a large number of endogenous binding partners for both Ezrin and Actin. Further, C-terminal actin binding capacity of the full length Ezrin is naturally shielded by its N-terminal, and only rendered active in the presence of Phosphatidylinositol bisphosphate (PIP2) or phosphorylation at the C-terminal threonine. Here, we demonstrate a strategy for the design, expression and purification of constructs, combining the Ezrin C-terminal actin binding domain, with functional elements such as fusion tags and fluorescence tags to facilitate purification and fluorescence microscopy based studies. For the first time, internal His tag was employed for purification of Ezrin actin binding domain based on in-silico modeling. The functionality (Ezrin-actin interaction) of these constructs was successfully demonstrated by using Total Internal Reflection Fluorescence Microscopy. This design can be extended to other members of the ERM family as well.

}, keywords = {Actins, Animals, Avian Proteins, Chickens, Cytoskeletal Proteins, In Vitro Techniques, Microfilament Proteins, Models, Molecular, Protein Interaction Domains and Motifs, Recombinant Fusion Proteins}, issn = {1932-6203}, doi = {10.1371/journal.pone.0123428}, author = {Shrivastava, Rohini and K{\"o}ster, Darius and Kalme, Sheetal and Mayor, Satyajit and Neerathilingam, Muniasamy} } @article {504, title = {Thioaptamers targeting dengue virus type-2 envelope protein domain III. [Protein Technology Core]}, journal = {Biochem Biophys Res Commun}, volume = {453}, year = {2014}, month = {2014 Oct 24}, pages = {309-15}, abstract = {

Thioaptamers targeting the dengue-2 virus (DENV-2) envelope protein domain III (EDIII) were developed. EDIII, which contains epitopes for binding neutralizing antibodies, is the putative host-receptor binding domain and is thus an attractive target for development of vaccines, anti-viral therapeutic and diagnostic agents. Thioaptamer DENTA-1 bound to DENV-2 EDIII adjacent to a known neutralizing antibody binding site with a dissociation constant of 154nM.

}, keywords = {Antibodies, Neutralizing, Antiviral Agents, Aptamers, Nucleotide, Base Sequence, Dengue Virus, Magnetic Resonance Spectroscopy, Viral Envelope Proteins}, issn = {1090-2104}, doi = {10.1016/j.bbrc.2014.09.053}, author = {Gandham, Sai Hari A and Volk, David E and Lokesh, Ganesh L R and Neerathilingam, Muniasamy and Gorenstein, David G} } @article {517, title = {Two acidic, anticoagulant PLA2 isoenzymes purified from the venom of monocled cobra Naja kaouthia exhibit different potency to inhibit thrombin and factor Xa via phospholipids independent, non-enzymatic mechanism. (Mass Spectrometry - Proteomics)}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e101334}, abstract = {

BACKGROUND: The monocled cobra (Naja kaouthia) is responsible for snakebite fatality in Indian subcontinent and in south-western China. Phospholipase A2 (PLA2; EC 3.1.1.4) is one of the toxic components of snake venom. The present study explores the mechanism and rationale(s) for the differences in anticoagulant potency of two acidic PLA2 isoenzymes, Nk-PLA2α (13463.91 Da) and Nk-PLA2β (13282.38 Da) purified from the venom of N. kaouthia.

PRINCIPAL FINDINGS: By LC-MS/MS analysis, these PLA2s showed highest similarity (23.5\% sequence coverage) with PLA2 III isolated from monocled cobra venom. The catalytic activity of Nk-PLA2β exceeds that of Nk-PLA2α. Heparin differentially regulated the catalytic and anticoagulant activities of these Nk-PLA2 isoenzymes. The anticoagulant potency of Nk-PLA2α was comparable to commercial anticoagulants warfarin, and heparin/antithrombin-III albeit Nk-PLA2β demonstrated highest anticoagulant activity. The anticoagulant action of these PLA2s was partially contributed by a small but specific hydrolysis of plasma phospholipids. The strong anticoagulant effect of Nk-PLA2α and Nk-PLA2β was achieved via preferential, non-enzymatic inhibition of FXa (Ki = 43 nM) and thrombin (Ki = 8.3 nM), respectively. Kinetics study suggests that the Nk-PLA2 isoenzymes inhibit their "pharmacological target(s)" by uncompetitive mechanism without the requirement of phospholipids/Ca(2+). The anticoagulant potency of Nk-PLA2β which is higher than that of Nk-PLA2α is corroborated by its superior catalytic activity, its higher capacity for binding to phosphatidylcholine, and its greater strength of thrombin inhibition. These PLA2 isoenzymes thus have evolved to affect haemostasis by different mechanisms. The Nk-PLA2β partially inhibited the thrombin-induced aggregation of mammalian platelets suggesting its therapeutic application in the prevention of unwanted clot formation.

CONCLUSION/SIGNIFICANCE: In order to develop peptide-based superior anticoagulant therapeutics, future application of Nk-PLA2α and Nk-PLA2β for the treatment and/or prevention of cardiovascular disorders are proposed.

}, keywords = {Anticoagulants, Blood Platelets, Chemical Fractionation, Chromatography, Liquid, Cobra Venoms, Factor Xa, Factor Xa Inhibitors, Isoenzymes, Kinetics, Phospholipases A2, Phospholipids, Sequence Analysis, Protein, Tandem Mass Spectrometry, Thrombin}, issn = {1932-6203}, doi = {10.1371/journal.pone.0101334}, author = {Mukherjee, Ashis K and Kalita, Bhargab and Thakur, Rupamoni} } @article {1009, title = {Tender coconut water an economical growth medium for the production of recombinant proteins in Escherichia coli. [Protein Technology Facility]}, journal = {BMC Biotechnol}, volume = {13}, year = {2013}, month = {2013 Sep 02}, pages = {70}, abstract = {

BACKGROUND: Escherichia coli is most widely used prokaryotic expression system for the production of recombinant proteins. Several strategies have been employed for expressing recombinant proteins in E.coli. This includes the development of novel host systems, expression vectors and cost effective media. In this study, we exploit tender coconut water (TCW) as a natural and cheaper growth medium for E.coli and Pichia pastoris.

RESULT: E.coli and P.pastoris were cultivated in TCW and the growth rate was monitored by measuring optical density at 600 nm (OD(600nm)), where 1.55 for E.coli and 8.7 for P.pastoris was obtained after 12 and 60 hours, respectively. However, variation in growth rate was observed among TCW when collected from different localities (0.15-2.5 at OD(600nm)), which is attributed to the varying chemical profile among samples. In this regard, we attempted the supplementation of TCW with different carbon and nitrogen sources to attain consistency in growth rate. Here, supplementation of TCW with 25 mM ammonium sulphate (TCW-S) was noted efficient for the normalization of inconsistency, which further increased the biomass of E.coli by 2 to 10 folds, and 1.5 to 2 fold in P.pastoris. These results indicate that nitrogen source is the major limiting factor for growth. This was supported by total nitrogen and carbon estimation where, nitrogen varies from 20 to 60 mg/100 ml while carbohydrates showed no considerable variation (2.32 to 3.96 g/100 ml). In this study, we also employed TCW as an expression media for recombinant proteins by demonstrating successful expression of maltose binding protein (MBP), MBP-TEV protease fusion and a photo switchable fluorescent protein (mEos2) using TCW and the expression level was found to be equivalent to Luria Broth (LB).

CONCLUSION: This study highlights the possible application of TCW-S as a media for cultivation of a variety of microorganisms and recombinant protein expression.

}, keywords = {Ammonium Sulfate, Biomass, Carbohydrates, Carbon, Cocos, Culture Media, Escherichia coli, Gene Expression, Nitrogen, Pichia, Recombinant Proteins}, issn = {1472-6750}, doi = {10.1186/1472-6750-13-70}, author = {Sekar, Narendrakumar and Veetil, Soumya Kariyadan and Neerathilingam, Muniasamy} }