Whole genome based identification of BAHD acyltransferase gene involved in piperine biosynthetic pathway in black pepper.

Black pepper (Piper nigrum L.), a crop of the genus Piper, is an important spice that has both economic and ecological significance. It is widely regarded as the "King of Spices" because of its pungency, attributed to the presence of piperine. BAHD acyl transferase, the crucial enzyme involved in the final step in piperine biosynthesis was the focus of our study and the aim was to identify the candidate isoform involved in biosynthesis of piperine. Reference genome-based analysis of black pepper identified six BAHD-AT isoforms and mapping of these sequences revealed that the isoforms were situated on six distinct chromosomes. By using specific primers for each of these transcripts, qPCR analysis was done in different tissues as well as berry stages to obtain detectable amplification products. Expression profiles of isoforms from chromosome 6 correlated well with piperine content compared to other five isoforms, across tissues and was therefore assumed to be involved in biosynthesis of piperine. In addition to this, we could also identify the binding sites of MYB transcription factor in the cis-regulatory regions of the isoforms. We also used in-silico docking and molecular dynamics simulation to calculate the binding free energy of the ligand and confirmed that among all the isoforms, BAHD-AT from chromosome 6 had the lowest free binding energy and highest affinity towards the ligand. Our findings are expected to aid the identification of new genes connecting enzymes involved in the biosynthetic pathway of piperine, which will have major implications for future research in metabolic engineering.Communicated by Ramaswamy H. Sarma.

In silico mutation of aromatic with aliphatic amino acid residues in Clostridium perfringens epsilon toxin (ETX) reduces its binding efficiency to Caprine Myelin and lymphocyte (MAL) protein receptors.

Enterotoxaemia (ET) is a severe disease that affects domestic ruminants, including sheep and goats, and is caused by Clostridium perfringens type B and D strains. The disease is characterized by the production of Epsilon toxin (ETX), which has a significant impact on the farming industry due to its high lethality. The binding of ETX to the host cell receptor is crucial, but still poorly understood. Therefore, the structural features of goat Myelin and lymphocytic (MAL) protein were investigated and defined in this study. We induced the mutations in aromatic amino acid residues of ETX and substituted them with aliphatic residues at domains I and II. Subsequently, protein-protein interactions (PPI) were performed between ETX (wild)-MAL and ETX (mutated)-MAL protein predicting the domain sites of ETX structure. Further, molecular dynamics (MD) simulation studies were performed for both complexes to investigate the dynamic behavior of the proteins. The binding efficiency between 'ETX (wild)-MAL protein' and 'ETX (mutated)-MAL protein complex' interactions were compared and showed that the former had stronger interactions and binding efficiency due to the higher stability of the complex. The MD analysis showed destabilization and higher fluctuations in the PPI of the mutated heterodimeric ETX-MAL complex which is otherwise essential for its functional conformation. Such kind of interactions with mutated functional domains of ligands provided much-needed clarity in understanding the pre-pore complex formation of epsilon toxin with the MAL protein receptor of goats. The findings from this study would provide an impetus for designing a novel vaccine for Enterotoxaemia in goats.Communicated by Ramaswamy H. Sarma.

Genome-wide identification and characterization of tissue specific long non-coding RNAs and circular RNAs in common carp (Cyprinus carpio L.).

Cyprinus carpio is regarded as a substitute vertebrate fish model for zebrafish. A varied category of non-coding RNAs is comprised of long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs). These ncRNAs were once considered non-functional "junk DNA" but research now shows they play important roles in gene expression regulation, chromatin modification, and epigenetic regulation. The systemic tissue-specific research of the lncRNAs and circRNAs of C. carpio is yet unexplored. A total of 468 raw RNA-Seq dataset across 28 distinct tissues from different varieties of common carp retrieved from public domain were pre-processing, mapped and assembled for lncRNA identification/ classification using various bioinformatics tools. A total of 33,990 lncRNAs were identified along with revelation of 9 miRNAs having 19 unique lncRNAs acting as their precursors. Additionally, 2,837 miRNAs were found to target 4,782 distinct lncRNAs in the lncRNA-miRNA-mRNA interaction network analysis, which resulted in the involvement of 3,718 mRNAs in common carp. A total of 22,854 circRNAs were identified tissue-wise across all the 28 tissues. Moreover, the examination of the circRNA-miRNA-mRNA interaction network revealed that 15,731 circRNAs were targeted by 5,906 distinct miRNAs, which in turn targeted 4,524 mRNAs in common carp. Significant signaling pathways like necroptosis, NOD-like receptor signaling pathway, hypertrophic cardiomyopathy, small cell lung cancer, MAPK signaling pathway, etc. were identified using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes. The web resource of common carp ncRNAs, named CCncRNAdb and available at http://backlin.cabgrid.res.in/ccncrnadb/ gives a comprehensive information about common carp lncRNAs, circRNAs, and ceRNAs interactions, which can aid in investigating their functional roles for its management.

Genome-wide identification and characterization of tissue-specific non-coding RNAs in black pepper (Piper nigrum L.).

Long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) are the two classes of non-coding RNAs (ncRNAs) present predominantly in plant cells and have various gene regulatory functions at pre- and post-transcriptional levels. Previously deemed as "junk", these ncRNAs have now been reported to be an important player in gene expression regulation, especially in stress conditions in many plant species. Black pepper, scientifically known as Piper nigrum L., despite being one of the most economically important spice crops, lacks studies related to these ncRNAs. From a panel of 53 RNA-Seq datasets of black pepper from six tissues, namely, flower, fruit, leaf, panicle, root, and stem of six black pepper cultivars, covering eight BioProjects across four countries, we identified and characterized a total of 6406 lncRNAs. Further downstream analysis inferred that these lncRNAs regulated 781 black pepper genes/gene products via miRNA-lncRNA-mRNA network interactions, thus working as competitive endogenous RNAs (ceRNAs). The interactions may be various mechanisms like miRNA-mediated gene silencing or lncRNAs acting as endogenous target mimics (eTMs) of the miRNAs. A total of 35 lncRNAs were also identified to be potential precursors of 94 miRNAs after being acted upon by endonucleases like Drosha and Dicer. Tissue-wise transcriptome analysis revealed 4621 circRNAs. Further, miRNA-circRNA-mRNA network analysis showed 432 circRNAs combining with 619 miRNAs and competing for the binding sites on 744 mRNAs in different black pepper tissues. These findings can help researchers to get a better insight to the yield regulation and responses to stress in black pepper in endeavor of higher production and improved breeding programs in black pepper varieties.

Unveiling the Wheat Microbiome under Varied Agricultural Field Conditions.

Wheat being the important staple food crop plays a significant role in nutritional security. A wide variety of microbial communities beneficial to plants and contributing to plant health and production are found in the rhizosphere. The wheat microbiome encompasses an extensive variety of microbial species playing a key role in sustaining the physiology of the crop, nutrient uptake, and biotic/abiotic stress resilience. This report presents wheat microbiome analysis under six different farm practices, namely, organic (Org), timely sown (TS), wheat after pulse crop (WAPC), temperature-controlled phenotyping facility (TCPF), maize-wheat cropping system (MW), and residue burnt field (Bur), using 16S rRNA sequencing methodology. The soil samples collected from either side of the wheat row were mixed to get a final sample set for DNA extraction under each condition. After the data preprocessing, microbial community analysis was performed, followed by functional analysis and annotation. An abundance of the phylum Proteobacteria was observed, followed by Acidobacteria, Actinobacteria, and Gemmatimonadetes in the majority of the samples, while relative abundance was found to vary at the genus level. Analysis against the Carbohydrate-Active Enzymes (CAZy) database showed a high number of glycoside hydrolase genes in the TS, TCPF, and WAPC samples, while the Org, MW, and Bur samples predominantly had glycosyltransferase genes and carbohydrate esterase genes were in the lowest numbers. Also, the Org and TCPF samples showed lower diversity, while rare and abundant species ranged from 12 to 25% and 20 to 32% of the total bacterial species in all the sets, respectively. These variations indicate that the different cropping sequence had a significant impact on soil microbial diversity and community composition, which characterizes its economic and environmental value as a sustainable agricultural approach to maintaining food security and ecosystem health. IMPORTANCE This investigation examined the wheat microbiome under six different agricultural field conditions to understand the role of cropping pattern on soil microbial diversity. This study also elaborated the community composition, which has importance in economic (role of beneficial community leading to higher production) and environmental (role of microbial diversity/community in safeguarding the soil health, etc.) arenas. This could lead to a sustainable farming approach for food security and improved ecosystem health. Also, the majority of the microbes are unculturable; hence, technology-based microcultivation will be a potential approach for harnessing other cultured microorganisms, leading to unique species for commercial production. The outcome of this research-accelerated work can provide an idea to the scientists/breeders/agronomists/pathologists under the mentioned field conditions regarding their influence over their crops.

Whole-Genome-Based Web Genomic Resource for Water Buffalo (Bubalus bubalis).

Water buffalo (Bubalus bubalis), belonging to the Bovidae family, is an economically important animal as it is the major source of milk, meat, and drought in numerous countries. It is mainly distributed in tropical and subtropical regions with a global population of approximately 202 million. The advent of low cost and rapid sequencing technologies has opened a new vista for global buffalo researchers. In this study, we utilized the genomic data of five commercially important buffalo breeds, distributed globally, namely, Mediterranean, Egyptian, Bangladesh, Jaffrarabadi, and Murrah. Since there is no whole-genome sequence analysis of these five distinct buffalo breeds, which represent a highly diverse ecosystem, we made an attempt for the same. We report the first comprehensive, holistic, and user-friendly web genomic resource of buffalo (BuffGR) accessible at http://backlin.cabgrid.res.in/buffgr/, that catalogues 6028881 SNPs and 613403 InDels extracted from a set of 31 buffalo tissues. We found a total of 7727122 SNPs and 634124 InDels distributed in four breeds of buffalo (Murrah, Bangladesh, Jaffarabadi, and Egyptian) with reference to the Mediterranean breed. It also houses 4504691 SSR markers from all the breeds along with 1458 unique circRNAs, 37712 lncRNAs, and 938 miRNAs. This comprehensive web resource can be widely used by buffalo researchers across the globe for use of markers in marker trait association, genetic diversity among the different breeds of buffalo, use of ncRNAs as regulatory molecules, post-transcriptional regulations, and role in various diseases/stresses. These SNPs and InDelscan also be used as biomarkers to address adulteration and traceability. This resource can also be useful in buffalo improvement programs and disease/breed management.

Delineation of molecular interactions of plant growth promoting bacteria induced ß-1,3-glucanases and guanosine triphosphate ligand for antifungal response in rice: a molecular dynamics approach.

BACKGROUND: The plant growth is influenced by multiple interactions with biotic (microbial) and abiotic components in their surroundings. These microbial interactions have both positive and negative effects on plant. Plant growth promoting bacterial (PGPR) interaction could result in positive growth under normal as well as in stress conditions. METHODS: Here, we have screened two PGPR's and determined their potential in induction of specific gene in host plant to overcome the adverse effect of biotic stress caused by Magnaporthe grisea, a fungal pathogen that cause blast in rice. We demonstrated the glucanase protein mode of action by performing comparative modeling and molecular docking of guanosine triphosphate (GTP) ligand with the protein. Besides, molecular dynamic simulations have been performed to understand the behavior of the glucanase-GTP complex. RESULTS: The results clearly showed that selected PGPR was better able to induce modification in host plant at morphological, biochemical, physiological and molecular level by activating the expression of ß-1,3-glucanases gene in infected host plant. The docking results indicated that Tyr75, Arg256, Gly258, and Ser223 of glucanase formed four crucial hydrogen bonds with the GTP, while, only Val220 found to form hydrophobic contact with ligand. CONCLUSIONS: The PGPR able to induce ß-1,3-glucanases gene in host plant upon pathogenic interaction and ß-1,3-glucanases form complex with GTP by hydrophilic interaction for induction of defense cascade for acquiring resistance against Magnaporthe grisea.

DeepAProt: Deep learning based abiotic stress protein sequence classification and identification tool in cereals.

The impact of climate change has been alarming for the crop growth. The extreme weather conditions can stress the crops and reduce the yield of major crops belonging to Poaceae family too, that sustains 50% of the world's food calorie and 20% of protein intake. Computational approaches, such as artificial intelligence-based techniques have become the forefront of prediction-based data interpretation and plant stress responses. In this study, we proposed a novel activation function, namely, Gaussian Error Linear Unit with Sigmoid (SIELU) which was implemented in the development of a Deep Learning (DL) model along with other hyper parameters for classification of unknown abiotic stress protein sequences from crops of Poaceae family. To develop this models, data pertaining to four different abiotic stress (namely, cold, drought, heat and salinity) responsive proteins of the crops belonging to poaceae family were retrieved from public domain. It was observed that efficiency of the DL models with our proposed novel SIELU activation function outperformed the models as compared to GeLU activation function, SVM and RF with 95.11%, 80.78%, 94.97%, and 81.69% accuracy for cold, drought, heat and salinity, respectively. Also, a web-based tool, named DeepAProt (http://login1.cabgrid.res.in:5500/) was developed using flask API, along with its mobile app. This server/App will provide researchers a convenient tool, which is rapid and economical in identification of proteins for abiotic stress management in crops Poaceae family, in endeavour of higher production for food security and combating hunger, ensuring UN SDG goal 2.0.

Revelation of candidate genes and molecular mechanism of reproductive seasonality in female rohu (Labeo rohita Ham.) by RNA sequencing.

BACKGROUND: Carp fish, rohu (Labeo rohita Ham.) is important freshwater aquaculture species of South-East Asia having seasonal reproductive rhythm. There is no holistic study at transcriptome level revealing key candidate genes involved in such circannual rhythm regulated by biological clock genes (BCGs). Seasonality manifestation has two contrasting phases of reproduction, i.e., post-spawning resting and initiation of gonadal activity appropriate for revealing the associated candidate genes. It can be deciphered by RNA sequencing of tissues involved in BPGL (Brain-Pituitary-Gonad-Liver) axis controlling seasonality. How far such BCGs of this fish are evolutionarily conserved across different phyla is unknown. Such study can be of further use to enhance fish productivity as seasonality restricts seed production beyond monsoon season. RESULT: A total of ~ 150 Gb of transcriptomic data of four tissues viz., BPGL were generated using Illumina TruSeq. De-novo assembled BPGL tissues revealed 75,554 differentially expressed transcripts, 115,534 SSRs, 65,584 SNPs, 514 pathways, 5379 transcription factors, 187 mature miRNA which regulates candidate genes represented by 1576 differentially expressed transcripts are available in the form of web-genomic resources. Findings were validated by qPCR. This is the first report in carp fish having 32 BCGs, found widely conserved in fish, amphibian, reptile, birds, prototheria, marsupials and placental mammals. This is due to universal mechanism of rhythmicity in response to environment and earth rotation having adaptive and reproductive significance. CONCLUSION: This study elucidates evolutionary conserved mechanism of photo-periodism sensing, neuroendocrine secretion, metabolism and yolk synthesis in liver, gonadal maturation, muscular growth with sensory and auditory perception in this fish. Study reveals fish as a good model for research on biological clock besides its relevance in reproductive efficiency enhancement.

De novo transcriptome sequencing assisted identification of terpene synthases from black pepper (Piper nigrum) berry.

Though the volatile profiles of black pepper have been reported already, the information on terpene synthase family genes is not known. In this study, using a combinatorial approach, the berry hybrid transcriptome assembly of llumina and nanopore sequencing, the entire terpene synthase family responsible for the biosynthesis of the flavor-imparting volatiles in black pepper berries was profiled. The profile shows 98 terpene synthases from various terpene synthesis pathways. Three important monoterpene synthases were also validated by targeted amplification, sequencing and homology modeling. This study provides the first of its kind information on the terpene synthase family profile in Piper nigrum, which is potentially a major step for further characterization of the functional terpene synthase genes in black pepper. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-00986-4.

Genome-Wide Association Study and Post-genome-Wide Association Study Analysis for Spike Fertility and Yield Related Traits in Bread Wheat.

Spike fertility and associated traits are key factors in deciding the grain yield potential of wheat. Genome-wide association study (GWAS) interwoven with advanced post-GWAS analysis such as a genotype-phenotype network (geno-pheno network) for spike fertility, grain yield, and associated traits allow to identify of novel genomic regions and represents attractive targets for future marker-assisted wheat improvement programs. In this study, GWAS was performed on 200 diverse wheat genotypes using Breeders' 35K Axiom array that led to the identification of 255 significant marker-trait associations (MTAs) (-log10P ≥ 3) for 15 metric traits phenotyped over three consecutive years. MTAs detected on chromosomes 3A, 3D, 5B, and 6A were most promising for spike fertility, grain yield, and associated traits. Furthermore, the geno-pheno network prioritised 11 significant MTAs that can be utilised as a minimal marker system for improving spike fertility and yield traits. In total, 119 MTAs were linked to 81 candidate genes encoding different types of functional proteins involved in various key pathways that affect the studied traits either way. Twenty-two novel loci were identified in present GWAS, twelve of which overlapped by candidate genes. These results were further validated by the gene expression analysis, Knetminer, and protein modelling. MTAs identified from this study hold promise for improving yield and related traits in wheat for continued genetic gain and in rapidly evolving artificial intelligence (AI) tools to apply in the breeding program.

Revealing Alteration in the Hepatic Glucose Metabolism of Genetically Improved Carp, Jayanti Rohu Labeo rohita Fed a High Carbohydrate Diet Using Transcriptome Sequencing.

Although feed cost is the greatest concern in aquaculture, the inclusion of carbohydrates in the fish diet, and their assimilation, are still not well understood in aquaculture species. We identified molecular events that occur due to the inclusion of high carbohydrate levels in the diets of genetically improved 'Jayanti rohu' Labeo rohita. To reveal transcriptional changes in the liver of rohu, a feeding experiment was conducted with three doses of gelatinized starch (20% (control), 40%, and 60%). Transcriptome sequencing revealed totals of 15,232 (4464 up- and 4343 down-regulated) and 15,360 (4478 up- and 4171 down-regulated) differentially expressed genes. Up-regulated transcripts associated with glucose metabolisms, such as hexokinase, PHK, glycogen synthase and PGK, were found in fish fed diets with high starch levels. Interestingly, a de novo lipogenesis mechanism was found to be enriched in the livers of treated fish due to up-regulated transcripts such as FAS, ACCα, and PPARγ. The insulin signaling pathways with enriched PPAR and mTOR were identified by Kyoto Encyclopedia of Genes and Genome (KEGG) as a result of high carbohydrates. This work revealed for the first time the atypical regulation transcripts associated with glucose metabolism and lipogenesis in the livers of Jayanti rohu due to the inclusion of high carbohydrate levels in the diet. This study also encourages the exploration of early nutritional programming for enhancing glucose efficiency in carp species, for sustainable and cost-effective aquaculture production.

Characterization of genetic diversity and population structure in wheat using array based SNP markers.

Genetic diversity is crucial for successful adaptation and sustained improvement in crops. India is bestowed with diverse agro-climatic conditions which makes it rich in wheat germplasm adapted to various niches. Germplasm repository consists of local landraces, trait specific genetic stocks including introgressions from wild relatives, exotic collections, released varieties, and improved germplasm. Characterization of genetic diversity is done using morpho-physiological characters as well as by analyzing variations at DNA level. However, there are not many reports on array based high throughput SNP markers having characteristics of genome wide coverage employed in Indian spring wheat germplasm. Amongst wheat SNP arrays, 35K Axiom Wheat Breeder's Array has the highest SNP polymorphism efficiency suitable for genetic mapping and genetic diversity characterization. Therefore, genotyping was done using 35K in 483 wheat genotypes resulting in 14,650 quality filtered SNPs, that were distributed across the B (~ 50%), A (~ 39%), and D (~ 10%) genomes. The total genetic distance coverage was 4477.85 cM with 3.27 SNP/cM and 0.49 cM/SNP as average marker density and average inter-marker distance, respectively. The PIC ranged from 0.09 to 0.38 with an average of 0.29 across genomes. Population structure and Principal Coordinate Analysis resulted in two subpopulations (SP1 and SP2). The analysis of molecular variance revealed the genetic variation of 2% among and 98% within subpopulations indicating high gene flow between SP1 and SP2. The subpopulation SP2 showed high level of genetic diversity based on genetic diversity indices viz. Shannon's information index (I) = 0.648, expected heterozygosity (He) = 0.456 and unbiased expected heterozygosity (uHe) = 0.456. To the best of our knowledge, this study is the first to include the largest set of Indian wheat genotypes studied exclusively for genetic diversity. These findings may serve as a potential source for the identification of uncharacterized QTL/gene using genome wide association studies and marker assisted selection in wheat breeding programs.

Revealing liver specific microRNAs linked with carbohydrate metabolism of farmed carp, Labeo rohita (Hamilton, 1822).

The role of microRNA in gene regulation during developmental biology has been well depicted in several organisms. The present study was performed to investigate miRNAs role in the liver tissues during carbohydrate metabolism and their targets in the farmed carp rohu, Labeo rohita, which is economically important species in aquaculture. Using Illumina-HiSeq technology, a total of 22,612,316; 44,316,046 and 13,338,434 clean reads were obtained from three small-RNA libraries. We have identified 138 conserved and 161 novel miRNAs and studies revealed that miR-22, miR-122, miR-365, miR-200, and miR-146 are involved in carbohydrate metabolism. Further analysis depicted mature miRNA and their predicted target sites in genes that were involved in developmental biology, cellular activities, transportation, etc. This is the first report of the presence of miRNAs in liver tissue of rohu and their comparative profile linked with metabolism serves as a vital resource as a biomarker.

Plant virus interaction mechanism and associated pathways in mosaic disease of small cardamom (Elettaria cardamomum Maton) by RNA-Seq approach.

Small cardamom (Elettaria cardamomum), grown in limited coastal tropical countries is one of the costliest and widely exported agri-produce having global turnover of >10 billion USD. Mosaic/marble disease is one of the major impediments that requires understanding of disease at molecular level. Neither whole genome sequence nor any genomic resources are available, thus RNA seq approach can be a rapid and economical alternative. De novo transcriptome assembly was done with Illumina Hiseq data. A total of 5317 DEGs, 2267 TFs, 114 pathways and 175,952 genic region putative markers were obtained. Gene regulatory network analysis deciphered molecular events involved in marble disease. This is the first transcriptomic report revealing disease mechanism mediated by perturbation in auxin homeostasis and ethylene signalling leading to senescence. The web-genomic resource (SCMVTDb) catalogues putative molecular markers, candidate genes and transcript information. SCMVTDb can be used in germplasm improvement against mosaic disease in endeavour of small cardamom productivity. Availability of genomic resource, SCMVTDb: http://webtom.cabgrid.res.in/scmvtdb/.

RNAseq analysis reveals drought-responsive molecular pathways with candidate genes and putative molecular markers in root tissue of wheat.

Drought is one of the major impediments in wheat productivity. Traditional breeding and marker assisted QTL introgression had limited success. Available wheat genomic and RNA-seq data can decipher novel drought tolerance mechanisms with putative candidate gene and marker discovery. Drought is first sensed by root tissue but limited information is available about how roots respond to drought stress. In this view, two contrasting genotypes, namely, NI5439 41 (drought tolerant) and WL711 (drought susceptible) were used to generate ~78.2 GB data for the responses of wheat roots to drought. A total of 45139 DEGs, 13820 TF, 288 miRNAs, 640 pathways and 435829 putative markers were obtained. Study reveals use of such data in QTL to QTN refinement by analysis on two model drought-responsive QTLs on chromosome 3B in wheat roots possessing 18 differentially regulated genes with 190 sequence variants (173 SNPs and 17 InDels). Gene regulatory networks showed 69 hub-genes integrating ABA dependent and independent pathways controlling sensing of drought, root growth, uptake regulation, purine metabolism, thiamine metabolism and antibiotics pathways, stomatal closure and senescence. Eleven SSR markers were validated in a panel of 18 diverse wheat varieties. For effective future use of findings, web genomic resources were developed. We report RNA-Seq approach on wheat roots describing the drought response mechanisms under field drought conditions along with genomic resources, warranted in endeavour of wheat productivity.

Liver-Specific microRNA Identification in Farmed Carp, Labeo bata (Hamilton, 1822), Fed with Starch Diet Using High-Throughput Sequencing.

The liver is an important central organ, which controls carbohydrate metabolism through maintaining glucose homeostasis by a tightly regulated system of genes or enzymes. The microRNAs are small non-coding RNAs playing an important role in the regulation of genes associated with developmental biology, physiology, metabolism, etc. Thus, in this study, we have intended to detect liver-specific microRNAs in farmed carp, Labeo bata, upon being fed a diet with different levels of carbohydrates. Here, we have conducted the experiment for 45 days using fingerlings of farmed carp fed with 20% (control), 40%, and 60% gelatinized starch levels. The liver tissues were collected from each treatment and processed for RNA isolation, small RNA library preparation, and high-throughput sequencing using Illumina NexSeq500. Through sequencing, 15,779,417 reads in 20% CHO, 13,959,039 in 40% CHO, and 13,661,950 in 60% CHO reads were generated for control and treated fishes using three small RNA libraries. We have investigated 445 novel and 231 conserved microRNAs in 20%, 40%, and 60% carbohydrate (CHO), respectively, through computational analysis. The differential expression analysis of miRNAs was carried out between different treatments compared with control and this study depicted 117 known and 114 novel miRNA genes involved in carbohydrate metabolic pathways. Further, target prediction and gene ontology analysis revealed that miRNAs were involved in several pathways such as signaling pathway, G protein pathway, complement receptor-mediated pathway, dopamine receptor signaling pathway, epidermal growth factor pathway, and notch signaling pathway. The predicted miRNA sites in targeted genes were associated with cellular activities, developmental biology, DNA binding, Golgi apparatus, extracellular region, catalytic activity, MAPK cascade, etc. Overall, we have generated a vital resource of liver-specific miRNAs involved in metabolic gene regulation. These studies further will help develop miRNA inhibitors to study their role during carbohydrate metabolism in farmed carp.

VigSatDB: genome-wide microsatellite DNA marker database of three species of Vigna for germplasm characterization and improvement.

Genus Vigna represented by more than 100 species is a source of nutritious edible seeds and sprouts that are rich sources of protein and dietary supplements. It is further valuable because of therapeutic attributes due to its antioxidant and anti-diabetic properties. A highly diverse and an extremely ecological niche of different species can be valuable genomic resources for productivity enhancement. It is one of the most underutilized crops for food security and animal feeds. In spite of huge species diversity, only three species of Vigna have been sequenced; thus, there is a need for molecular markers for the remaining species. Computational approach of microsatellite marker discovery along with evaluation of polymorphism utilizing available genomic data of different genotypes can be a quick and an economical approach for genomic resource development. Cross-species transferability by e-PCR over available genomes can further prioritize the potential SSR markers, which could be used for genetic diversity and population differentiation of the remaining species saving cost and time. We present VigSatDB-the world's first comprehensive microsatellite database of genus Vigna, containing >875 K putative microsatellite markers with 772 354 simple and 103 865 compound markers mined from six genome assemblies of three Vigna species, namely, Vigna radiata (Mung bean), Vigna angularis (Adzuki bean) and Vigna unguiculata (Cowpea). It also contains 1976 validated published markers. Markers can be selected on the basis of chromosomes/location specificity, and primers can be generated using Primer3core tool integrated at backend. Efficacy of VigSatDB for microsatellite loci genotyping has been evaluated by 15 markers over a panel of 10 diverse genotype of V. radiata. Our web genomic resources can be used in diversity analysis, population and varietal differentiation, discovery of quantitative trait loci/genes, marker-assisted varietal improvement in endeavor of Vigna crop productivity and management.

Uncovering Genomic Regions Associated With 36 Agro-Morphological Traits in Indian Spring Wheat Using GWAS.

Wheat genetic improvement by integration of advanced genomic technologies is one way of improving productivity. To facilitate the breeding of economically important traits in wheat, SNP loci and underlying candidate genes associated with the 36 agro-morphological traits were studied in a diverse panel of 404 genotypes. By using Breeders' 35K Axiom array in a comprehensive genome-wide association study covering 4364.79 cM of the wheat genome and applying a compressed mixed linear model, a total of 146 SNPs (-log10 P ≥ 4) were found associated with 23 traits out of 36 traits studied explaining 3.7-47.0% of phenotypic variance. To reveal this a subset of 260 genotypes was characterized phenotypically for six quantitative traits [days to heading (DTH), days to maturity (DTM), plant height (PH), spike length (SL), awn length (Awn_L), and leaf length (Leaf_L)] under five environments. Gene annotations mined ∼38 putative candidate genes which were confirmed using tissue and stage specific gene expression data from RNA Seq. We observed strong co-localized loci for four traits (glume pubescence, SL, PH, and awn color) on chromosome 1B (24.64 cM) annotated five putative candidate genes. This study led to the discovery of hitherto unreported loci for some less explored traits (such as leaf sheath wax, awn attitude, and glume pubescence) besides the refined chromosomal regions of known loci associated with the traits. This study provides valuable information of the genetic loci and their potential genes underlying the traits such as awn characters which are being considered as important contributors toward yield enhancement.

Development of species specific putative miRNA and its target prediction tool in wheat (Triticum aestivum L.).

MicroRNA are 20-24 nt, non-coding, single stranded molecule regulating traits and stress response. Tissue and time specific expression limits its detection, thus is major challenge in their discovery. Wheat has limited 119 miRNAs in MiRBase due to limitation of conservation based methodology where old and new miRNA genes gets excluded. This is due to origin of hexaploid wheat by three successive hybridization, older AA, BB and younger DD subgenome. Species specific miRNA prediction (SMIRP concept) based on 152 thermodynamic features of training dataset using support vector machine learning approach has improved prediction accuracy to 97.7%. This has been implemented in TamiRPred ( http://webtom.cabgrid.res.in/tamirpred ). We also report highest number of putative miRNA genes (4464) of wheat from whole genome sequence populated in database developed in PHP and MySQL. TamiRPred has predicted 2092 (>45.10%) additional miRNA which was not predicted by miRLocator. Predicted miRNAs have been validated by miRBase, small RNA libraries, secondary structure, degradome dataset, star miRNA and binding sites in wheat coding region. This tool can accelerate miRNA polymorphism discovery to be used in wheat trait improvement. Since it predicts chromosome-wise miRNA genes with their respective physical location thus can be transferred using linked SSR markers. This prediction approach can be used as model even in other polyploid crops.