Identification, characterization, and comprehensive expression profiling of floral master regulators in pigeon pea (Cajanus cajan [L.] Millspaugh).

Pigeon pea is an important protein-rich pulse crop. Identification of flowering master regulators in pigeon pea is highly imperative as indeterminacy and late flowering are impediments towards yield improvement. A genome-wide analysis was performed to explore flowering orthologous groups in pigeon pea. Among the 412 floral orthologs identified in pigeon pea, 148 genes belong to the meristem identity, photoperiod-responsive, and circadian clock-associated ortholog groups. Our comparative genomics study revealed purifying selection pressures (ka/ks) on floral orthologs, and duplication patterns and evolution through synteny with other model species. Phylogenetic analysis of floral genes substantiated a connection between pigeon pea plant architecture and flowering time as all the PEBP domain-containing genes belong to meristem identity floral networks of pigeon pea. Expression profiling of eleven major orthologs in contrasting determinate and indeterminate genotypes indicated that these orthologs might be involved in flowering regulation. Expression of floral inducer, FT, and floral repressor, TFL1, was non-comparable in indeterminate genotypes across all the developmental stages of pigeon pea. However, dynamic FT/TFL1 expression ratio detected in all tissues of both the genotypes suggested their role in floral transition. One TFL1 ortholog having high sequence conserveness across pigeon pea genotypes showed differential expression indicating genotype-dependent regulation of this ortholog. Presence of conserved 6mA-methylation patterns in light-responsive elements and in other cis-regulatory elements of FT and TFL1 across different plant genotypes indicated possible involvement of epigenetic regulation in flowering.

Colonization potential of endophytes from halophytic plants growing in the "Runn of Kutch" salt marshes and their contribution to mitigating salt stress in tomato cultivation.

Increasing soil salinity depreciates the quantity of the crop produce. Looking at the tremendous potential of plant-associated microorganisms in salinity stress mitigation, it would be very useful in exploring and deciphering salt-tolerant microorganisms from halophytic plants and their utilization in cultivated plants. With this aim, in the present study, four halophytic plants were taken from Rann of Kutch, and bacterial endophytes were isolated from different plant organs. These endophytes were characterized by plant growth and health promotion features. The molecular identification was done based on 16 s rRNA sequence similarity. It was found that the endophytic bacteria isolated from 4 different halophytes found sharing phylogenetic relatedness. Four potential endophytes Alkalihalobacillus gibsonii 2H2, Achromobacter insuavis 2H18, Terribacillus halophilus 2H20, and Bacillus siamensis 4H1 were tested in tomato for salinity stress alleviation. Changes in the levels of antioxidants were analyzed. Total chlorophyll, total phenolics, malondialdehyde, and proline content indicated reduced damage in the plant system due to salinity by the application of endophytes. All the treatments exhibited low levels of electrolyte leakage. The accumulation of enzymatic reactive oxygen species scavengers was assessed from the levels of peroxidase, catalase, superoxide dismutase, phenylalanine ammonia-lyase, ascorbate peroxidase, and guiacol peroxidase. The NBT and DAB staining confirmed the findings. The reduction in the accumulation of Na+ ions in tomato leaves was visualized using Sodium Green probes under CSLM and found to be lowest in Terribacillus halophilus 2H20 and Bacillus siamensis 4H1 inoculated plants. The endophyte Terribacillus halophilus 2H20 was the most promising isolate. The colonization in tomato roots was confirmed using a cell tracker system. Results showed that the endophytes were found to have salinity stress mitigation traits. The efficiency could be further improved with the combination of other endophytes tested earlier.

Host-Delivered RNA Interference for Durable Pest Resistance in Plants: Advanced Methods, Challenges, and Applications.

Insect-pests infestation greatly affects global agricultural production and is projected to become more severe in upcoming years. There is concern about pesticide application being ineffective due to insect resistance and environmental toxicity. Reduced effectiveness of Bt toxins also made the scientific community shift toward alternative strategies to control devastating agricultural pests. With the advent of host-delivered RNA interference, also known as host-induced gene silencing, targeted insect genes have been suppressed through genetic engineering tools to deliver a novel insect-pest resistance strategy for combating a number of agricultural pests. This review recapitulates the possible mechanism of host-delivered RNA interference (HD-RNAi), in particular, the silencing of target genes of insect-pests. We emphasize the development of the latest strategies against evolving insect targets including designing of artificial microRNAs, vector constructs, and the benefit of using plastid transformation to transform target RNA-interfering genes. Advantages of using HD-RNAi over other small RNA delivery modes and also the supremacy of HD-RNAi over the CRISPR-Cas system particularly for insect resistance have been described. However, the broader application of this technology is restricted due to its several limitations. Using artificial miRNA designs, the host-delivered RNAi + Bt combinatorial approach and chloroplast transformation can overcome limitations of RNAi. With careful design and delivery approaches, RNAi promises to be extremely valuable and effective plant protection strategy to attain durable insect-pest resistance in crops. Development of transgenic plant using novel strategies to achieve durable resistance against the target insect.

Amelioration of cold-induced sweetening in potato by RNAi mediated silencing of StUGPase encoding UDP-glucose pyrophosphorylase.

Cold-induced sweetening (CIS) is an unwanted physiological phenomenon in which reducing sugars (RS) get accumulated in potato (Solanum tuberosum) upon cold storage. High RS content makes potato commercially unsuitable for processing due to the unacceptable brown color in processed products like chips, fries, etc., and the production of a potential carcinogen, acrylamide. UDP-glucose pyrophosphorylase (UGPase) catalyzes the synthesis of UDP-glucose towards the synthesis of sucrose and is also involved in the regulation of CIS in potato. The objective of the present work was RNAi-mediated downregulation of the StUGPase expression level in potato for the development of CIS tolerant potato. Hairpin RNA (hpRNA) gene construct was developed by placing UGPase cDNA fragment in sense and antisense orientation intervened by GBSS intron. Internodal stem explants (cv. Kufri Chipsona-4) were transformed with hpRNA gene construct, and 22 transgenic lines were obtained by PCR screening of putative transformants. Four transgenic lines showed the highest level of RS content reduction following 30 days of cold storage, with reductions in sucrose and RS (glucose & fructose) levels of up to 46% and 57.5%, respectively. Cold stored transgenic potato of these four lines produced acceptable chip colour upon processing. The selected transgenic lines carried two to five copies of the transgene. Northern hybridization revealed an accumulation of siRNA with a concomitant decrease in the StUGPase transcript level in these selected transgenic lines. The present work demonstrates the efficacy of StUGPase silencing in controlling CIS in potato, and the strategy can be employed for the development of CIS tolerant potato varieties.

ROS generated from biotic stress: Effects on plants and alleviation by endophytic microbes.

Aerobic living is thought to generate reactive oxygen species (ROS), which are an inevitable chemical component. They are produced exclusively in cellular compartments in aerobic metabolism involving significant energy transfer and are regarded as by-products. ROS have a significant role in plant response to pathogenic stress, but the pattern varies between necrotrophs and biotrophs. A fine-tuned systemic induction system is involved in ROS-mediated disease development in plants. In regulated concentrations, ROS act as a signaling molecule and activate different pathways to suppress the pathogens. However, an excess of these ROS is deleterious to the plant system. Along with altering cell structure, ROS cause a variety of physiological reactions in plants that lower plant yield. ROS also degrade proteins, enzymes, nucleic acids, and other substances. Plants have their own mechanisms to overcome excess ROS and maintain homeostasis. Microbes, especially endophytes, have been reported to maintain ROS homeostasis in both biotic and abiotic stresses by multiple mechanisms. Endophytes themselves produce antioxidant compounds and also induce host plant machinery to supplement ROS scavenging. The structured reviews on how endophytes play a role in ROS homeostasis under biotic stress were very meager, so an attempt was made to compile the recent developments in ROS homeostasis using endophytes. This review deals with ROS production, mechanisms involved in ROS signaling, host plant mechanisms in alleviating oxidative stress, and the roles of endophytes in maintaining ROS homeostasis under biotic stress.

Identification and characterization of chickpea genotypes for early flowering and higher seed germination through molecular markers.

BACKGROUND: Chickpea is the fourth most important legume crop contributing 15.42% to the total legume production and a rich source of proteins, minerals, and vitamins. Determination of genetic diversity of wild and elite cultivars coupled with early flowering and higher seed germination lines are quintessential for variety improvement. METHODS AND RESULTS: In the present study, we have analyzed the genetic diversity, population structure, cross-species transferability, and allelic richness in 50 chickpea collections using 23 Inter simple sequence repeats (ISSR) markers. The observed parameters such as allele number varied from 3 to 16, range of allele size varied from 150 to 1600 bp and polymorphic information content (PIC) range lies in between 0.15 and 0.49. Dendrogram was constructed with ISSR marker genotypic data and classified 50 chickpea germplasms into groups I and II, where the accession P 74 - 1 is in group I and the rest are in group II. Dendrogram, Principal component analysis (PCA), dissimilarity matrix, and Bayesian model-based genetic clustering of 50 chickpea germplasms revealed that P 74 - 1 and P 1883 are very diverse chickpea accessions. CONCLUSION: Based on genetic diversity analysis, 15 chickpea germplasm having been screened for early flowering and higher seed germination and found that the P 1857-1 and P 3971 have early flowering and higher seed germination percentage in comparison to P 1883 and other germplasm. These agronomic traits are essential for crop improvement and imply the potential of ISSR markers in crop improvement.

Surface sterilization for isolation of endophytes: Ensuring what (not) to grow.

Endophytic microbiota opens a magnificent arena of metabolites that served as a potential source of medicines for treating a variety of ailments and having prospective uses in agriculture, food, cosmetics, and many more. There are umpteen reports of endophytes improving the growth and tolerance of plants. In addition, endophytes from lifesaving drug-producing plants such as Taxus, Nothapodytes, Catharanthus, and so forth have the ability to produce host mimicking compounds. To harness these benefits, it is imperative to isolate the true endophytes, not the surface microflora. The foremost step in endophyte isolation is the removal of epiphytic microbes from plant tissues, called as surface sterilization. The success of surface sterilization decides "what to grow" (the endophytes) and "what not to grow" (the epiphytes). It is very crucial to use an appropriate sterilant solution, concentration, and exposure time to ensure thorough surface disinfection with minimal damage to the endophytic diversity. Commonly used surface sterilants include sodium hypochlorite (2%-10%), ethanol (70%-90%), mercuric chloride (0.1%), formaldehyde (40%), and so forth. In addition, the efficiency could further be improved by pretreatment with surfactants such as Triton X-100, Tween 80, and Tween 20. This review comprehensively deals with the various sterilants and sterilization methods for the isolation of endophytic microbes. In addition, the mechanisms and rationale behind using specific surface sterilants have also been elaborated at length.

Success of microbial genes based transgenic crops: Bt and beyond Bt.

Transgenic technology could hold the key to help farmers to fulfill the ever increasing fast-paced global demand for food. Microbes have always wondered us by their potentials and thriving abilities in the extreme conditions. The use of microorganisms as a gene source in transgenic development is a promising option for crop improvement. The aforesaid approach has already for improving the characteristics of food, industrial, horticulture, and floriculture crops. Many transgenic crops containing microbial genes have been accepted by the farmers and consumers worldwide over the last few decades. The acceptance has brought remarkable changes in the status of society by providing food safety, economic, and health benefits. Among transgenic plants harboring microbial genes, Bacillus thuringiensis (Bt) based transgenic were more focused and documented owing to its significant performance in controlling insects. However, other microbial gene-based transgenic plants have also reserved their places in the farmer's field globally. Therefore, in this review, we have thrown some light on successful transgenic plants harboring microbial genes other than Bt, having application in agriculture. Also, we presented the role of microbial genetic element and product thereof in the inception of biotechnology and discussed the potential of microbial genes in crop improvement.

Nitrogen fixing cereal: A rising hero towards meeting food security.

Nitrogen serves as one of the primary components of major biomolecules and thus extends a significant contribution to crop growth and yield. But the inability of plants to utilize freely available atmospheric N2 makes the whole agricultural system dependent on chemical fertilizers, which incur significant input cost to supplement required quantities of nitrogen to crops. Only bacteria and archaea have been gifted with the power of drawing free N2 from air to convert them into NH3, which is one of the two utilizable forms of nitrogen taken up by plants. Legumes, the only family of crops, can engage themselves in symbiotic nitrogen fixation where they establish a mutualistic relationship with nitrogen-fixing bacteria and in turn, can waive off the necessity of adding nitrogen fertilizers. Sincere effort, therefore, has been undertaken to incorporate this capability of nitrogen-fixation into non-legume crops, especially cereals which make up a vital portion in the food basket. Biotechnological interventions have also played important role in providing nitrogen fixing trait to non-legumes. This review takes up an effort to look into and accumulate all the important updates to date regarding nitrogen-fixing non-legumes with a special focus on cereals, which is one of the most important future goals in the field of science in the present era.