The main intent of northern blotting is an essential strategy in molecular biology to quantify the particular messenger RNA (mRNA). The very initial step is to identify the cells that express a specific gene, therefore providing a better understanding regarding the physiological role of the protein that has been encoded. Understanding the nutritional, hormonal, or environmental factors which governed by a particular gene is the primary aim of measuring mRNA. In addition to identifying transcript presence, it also provides size and a comparative analysis for transcript abundance over the single membrane. It's been effectively adapted in recent years to investigate and validate the expression in addition to size of short noncoding RNA. The denaturing agarose/polyacrylamide gel should be used to segregate RNA based on their size before it can be transferred on the solid surface in a way that maintains the topology distribution of the gel for the purpose of preparing northern blot to hybridization. This chapter clearly emphasizes the step-by-step protocol for northern blotting.

The COVID-19 pandemic has significantly reshaped various aspects of daily life, including the way we perceive and experience food. This review explores the emerging issue of taste alterations among individuals recovering from COVID-19, focusing on the changes in taste preferences and the broader consequences for culinary experiences. It investigates the complex relationship between SARS-CoV-2 and the chemosensory systems, highlighting the physiological mechanisms that contribute to disturbances in taste and smell. Beyond individual sensory changes, the discussion expands to consider the impact on the food industry, culinary traditions, and social dining habits. Through addressing these evolving sensory experiences, the review emphasizes the need for greater awareness and adaptive strategies for those affected. It also offers valuable perspectives for researchers, clinicians, and food professionals aiming to support recovery and innovation in the post-pandemic culinary landscape. Ultimately, this examination contributes to a deeper understanding of COVID-19’s lasting influence on human taste and gastronomy.

Taster categorisation uses bitter thiourea compounds like propylthiouracil (PROP) and phenylthiocarbamide (PTC), which are frequently associated with amino acid alterations at positions 49, 262 and 296 in human taste 2 receptor member 38 (hTAS2R38). Since the hTAS2R38 protein lacked a crystallographic structure, it was modelled using contact-guided iterative threading assembly refinement, its residues were mutated and refined, and the binding pocket area and volume were assessed using CASTp. Bitter thiourea molecules were docked using the ligand extra precision module and the receptor-ligand complex was manually positioned in a fully hydrated, equilibrated 1-palmitoyl-2-oleoylphosphatidylcholine bilayer using the CHARMM GUI membrane constructor, a 100 ns simulation was carried out using the Desmond program. Analysis revealed that the PROP binds to the allosteric hydrophobic pocket of hTAS2R38 and forms a hydrogen bond with ASN190. The native structure (hTAS2R38PAV) has a higher glide energy (-24.164 kcal/mol) and docking score (-7.212 kcal/mol) than mutants, corroborating our taste preference study. In contrast, PTC lacks hydrogen bonds in the binding pocket but exhibits pi-pi stacking interactions with the native structure. Structures with mutations at the 49th or 296th position showed the largest root mean square deviations and fluctuations. A triple mutation increases surface area and volume, making the 262nd position critical to the binding pocket. These results highlight the functional roles of these three residues in hTAS2R38.

Sustainable agriculture is severely affected by insect as well as pest attack. The manipulation of numerous genes to control pests by RNAi involves post-transcriptional gene silencing that is sequence-specific. This chapter is the first to describe the use of hairpin RNAi in tobacco plants as a novel approach to target insect pests and the potential impact this method could have on future pest management. Here we describe the generation of hpRNA constructs in tobacco for controlling insect pests. First, it describes how to make and test hpRNA constructs aimed at essential pest genes such as those required for development or survival. In this part of our study, the chapter on RNAi design explains in depth the procedures to create these RNAi vectors using cloning strategies in vector structure. It also deals with the use of Agrobacterium-mediated transformation for both tobacco plants and selection of transgenic plants. We look at approaches for measuring the impact of RNAi in these plants, such as gene expression analysis and evaluation of host resistance in laboratory or field conditions. This chapter is a comprehensive guide by giving an in-depth view on the steps from construct designing to in field applications; thus it will serve as reference for all future workers of RNAi technology looking towards effective and eco-friendly ways of pest management in agriculture.

Chloroplasts, originating from ancestral cyanobacteria, are essential plastids involved in photosynthesis and other key metabolic processes, making them vital components of plant cells. Chloroplast transformation is a preferred alternative to nuclear transformation in plant genetic engineering due to its numerous advantages and positive outcomes. This chapter explores the available various chloroplast transformation vectors, cloning the gene of interest (GOI) in the chloroplast transformation vector with step-by-step protocol, and preparation of the vector for chloroplast transformation of tobacco using leaf disc explants through a biolistic method for efficient gene delivery. Additionally, it highlights how crucial some factors are for the effective integration and expression of the chloroplast genome by DNA-coated particle bombardment, including microcarrier type, DNA concentration, helium accelerating pressure, leaf layout and firing distance, and duration of pretreatment for explants. Further explored the generation of homoplasmic plants through re-regeneration with antibiotic selection confirmed correct cassette insertion and homoplasmy using PCR and Southern blot analyses and verified maternal inheritance of chloroplast-transferred genes by examining green and albino seedlings from reciprocal crosses between control and transplastomic tobacco plants.

The most significant traits in agricultural biotechnology are those resulting from expression of transgenes, introduced to plant species such as cotton which has incorporated genes encoding resistance against biotic and abiotic stress. This chapter emphasizes on characterization of transgenic cotton plants in terms of polymerase chain reaction (PCR) techniques. This chapter describes in detail the protocol for primer design, genomic DNA and total RNA extraction, cDNA synthesis from RNA, master mix preparation, thermo cycler procedure, and finally analysis of PCR amplified product by agarose gel electrophoresis. This chapter describes not only the basic foundations of PCR, for instance, primer design, thermal cycling, and DNA amplification. It also covers the aspect of using reverse transcription PCR (RT-PCR) which is a type to measure gene expression amounts by transcribing RNA as complementary DNA (cDNA) before amplification. One needs to use RT-PCR for a more functional readout, especially if trying to measure the change caused by the transgene insertions.

In this chapter we focus for characterizing the RNA interference (RNAi) tobacco plants and detecting existence of the small interfering RNAs (siRNAs). RNAi, the most powerful gene silencing approach that engages in introducing double-stranded RNA (dsRNA) homologous to the target gene. The popular model plant system, tobacco, is employed for studying RNAi because of easy transformation as well as fast growth. This chapter emphasizes the steps that include total RNA extraction, cDNA synthesis, qRT-PCR amplification, as well as northern blotting. To segregate the DNA contamination from isolated total RNA extracted using TRIzol method, DNase treatment was carried out. PCR amplification was performed to check for the target gene and housekeeping gene. Northern blot experiment was done to examine the existence of siRNAs. The experiment worked in that RNAi-mediated gene silencing occurred in the tobacco plants, as proven by lower expression of the target gene and detection of siRNAs. This research offers a great protocol for the analysis of RNAi tobacco plants and the detection of siRNAs and can be utilized in many different research fields, such as plant biotechnology and functional genomics.

Transgenic cotton plants are responsible of a greater breakthrough in the field of agricultural biotechnology which focuses on improving plant characteristics such as pest resistance, drought tolerance, and fiber quality. The precise molecular characterization of these genetically transformed plants is fundamental in assessing the insertion, inheritance, and expression of transgenes. The β-glucuronidase (GUS) assay is among the conventional tools to test transgenic cotton that offer a consistent and productive process for determining gene expression. This chapter offers the exploration of the GUS assay using transgenic cotton plants, which may be useful for gene development. The enzyme that gives the blue product is β-glucuronidase, which in turn cleaves its substrate (5-bromo-4-chloro-3-indolyl β-D-glucuronide [X-Gluc]). It signals a change in color which can be seen directly as an indicator for the transgene expression. The histochemical assay significance is based on tissue fixation time, incubation periods, staining, and decolorizing of chlorophyll procedures for reproducible results of putative transgenic lines. It also covers approaches for optimizing assay conditions, such as changing the levels of substrates and incubation times to improve staining clarity and intensity. This chapter provides the step-by-step protocol for initial analysis of transgenic cotton by reporter assay by GUS histochemical staining.

Hypertension (HT) significantly impacts cardiovascular health (CVH) by exerting chronic stress on arteries and the heart, leading to severe health complications. This review explores the intricate relationship between dietary choices and high blood pressure (BP). Dietary choices are crucial in both the development and management of high BP. Excessive sodium intake is a well-documented risk factor for HT, and strategies to reduce salt consumption can mitigate its adverse effects. Processed and packaged foods also pose severe risks due to hidden sodium and other unhealthy ingredients, making healthier alternatives essential. Potassium plays a vital role in managing BP, with potassium-rich foods supporting a balanced diet. Saturated and trans fats contribute to elevated BP, and healthier fat alternatives are recommended. Excessive sugar consumption negatively affects BP, with hidden sugars in popular foods requiring practical strategies for reduction. Alcohol and caffeine also influence BP, and moderate consumption is advised to avoid potential risks. The Mediterranean diet offers a holistic, evidence-based approach to enhancing CVH, with practical advice and meal plans for a flavourful and health-promoting dietary regimen.

Genetically engineered plants are emerging as an increasingly popular tactics for combating insect pests, offering the possibility of increased crop yields with reduced reliance on toxic pesticides. A notable instance is the emergence of Bt-transgenic cotton that incorporates a gene obtained from the bacterium Bacillus thuringiensis which releases a toxin lethal for certain insect pests especially bollworms. This strategy has become extensively utilized because Bt-cotton has enabled for significant rises in cotton production. Concerns are being expressed regarding the possibility for insect resistance to Bt toxins along with the chance of adverse impacts to organisms that are not targets. This work seeks to provide an optimized manual approach for producing Bt-transgenic cotton cultivars that can successfully manage insect pests while reducing potential negative consequences. The main procedures for introducing genes, transforming plants, and choosing transgenic lines exhibiting desired characteristics will all be covered in the protocol. The resultant transgenic cotton will undergo extensive investigation and assessment in order to determine their overall agronomic performance, potential effects on nontarget organisms, and effectiveness against target pests. This research aims to actively contribute toward the ethical creation and implementation of insect-resistant genetically modified cotton by offering a strong and well-characterized manual process, balancing the need for improved agricultural output and preserving the environment. This manual provides a step by step protocol for successful transformation of transgenic cotton.

Nanopollution (NPOs), a burgeoning consequence of the widespread use of nanoparticles (NPs) across diverse industrial and consumer domains, has emerged as a critical environmental issue. While extensive research has scrutinized the repercussions of NPs pollution on ecosystems and human health, scant attention has been directed towards unraveling its implications for plant life. This comprehensive review aims to bridge this gap by delving into the nuanced interplay between NPOs and plant metabolism, encompassing both primary and secondary processes. Our exploration encompasses an in-depth analysis of the intricate mechanisms governing the interaction between plants and NPs. This involves a thorough examination of how physicochemical properties such as size, shape, and surface characteristics influence the uptake and translocation of NPs within plant tissues. The impact of NPOs on primary metabolic processes, including photosynthesis, respiration, nutrient uptake, and water transport. Additionally, this study explored the multifaceted alterations in secondary metabolism, shedding light on the synthesis and modulation of secondary metabolites in response to NPs exposure. In assessing the consequences of NPOs for plant life, we scrutinize the potential implications for plant growth, development, and environmental interactions. The intricate relationships revealed in this review underscore the need for a holistic understanding of the plant-NPs dynamics. As NPs become increasingly prevalent in ecosystems, this investigation establishes a fundamental guide that underscores the importance of additional research to shape sustainable environmental management strategies and address the extensive effects of NPs on the development of plant life and environmental interactions.

Genetic variations in taste receptors are associated with gustatory perception and obesity, which in turn affects dietary preferences. Given the increasing tendency of people with obesity choosing sweet, high-fat meals, the current study assessed the cross-regulation of two polymorphisms of the sweet taste receptor (T1R2/T1R3), rs35874116 and rs307355, on fat sensitivity in Indian adults. We investigated the association between taste sensitivity and BMI in the T1R2, T1R3, and CD36 polymorphic and non-polymorphic groups. The general labelled magnitude scale (gLMS) was used to assess the taste sensitivity of 249 participants in addition to anthropometric data. TaqMan Probe-based RT-PCR was employed to determine the polymorphisms. Additionally, the colorimetric method utilizing 3, 5-dinitro salicylic acid was used to evaluate the participants' salivary amylase activity. The mean detection thresholds for linoleic acid (LA) and sucrose were greater in individuals with obesity (i.e., 0.97 ± 0.08 mM and 0.22 ± 0.02 M, respectively) than in healthy adults (p < 0.0001), indicating lower sensitivity. Moreover, it was found that a greater proportion of persons with obesity fall into the polymorphic groups (i.e., 52

Gustin, a trophic factor for taste bud development, and its polymorphism at rs2274333 influence taste perception of 6-n-propylthiouracil (PROP) and fungiform papillae (FP) density. The PROP taster status affects dietary fat sensing and body composition. However, there is a paucity of research on the gustin genotype with dietary fat perception, PROP tasting ability, and body mass index (BMI). Thus, taste sensitivity to fat and bitterness was evaluated in 178 healthy individuals. The general labelled magnitude scale was used to determine suprathreshold taste intensity ratings, whereas the alternative forced choice approach was used to estimate the taste-sensing ability. The FP density was assessed by applying blue-coloured food dye over the anterior region of the tongue. Restriction fragment length polymorphism was used to detect the genetic polymorphism (rs2274333) in the CA-VI gene. Fisher's chi-square analysis showed that the CA-VI genotype and allelic frequencies significantly correlated (p<0.001) with the PROP taster status and BMI. Healthy individuals with AA genotypes of the CA-VI polymorphism and PROP super-tasters demonstrated stronger gustatory sensitivity for linoleic acid (LA) with greater FP density in comparison to individuals with AG/GG genotypes and other PROP taster groups. Stepwise forward multiple regression analysis indicates that BMI and PROP taster status significantly influence the LA sensing ability. The suprathreshold intensity rating for LA was also significantly impacted by PROP taster status and CA-VI genotypes, with a variation of 73.3%. Overall, our findings show a relationship between the taste papillae environment and the CA-VI genetic mutation at rs2274333, which influenced the gustatory preference for dietary fat and bitter taste.

Berberis tinctoria Lesch. is an endemic wild plant with significant medicinal value in traditional ethnomedicine. Although its chemical composition has not yet been studied, tribal communities in Nilgiri-Hills (NH) and Palani-Hills (PH) use it as food and to cure various ailments. The present study aims to analyze the influence of ecological factors on the accumulation of active compounds and antioxidants in B. tinctoria fruits. The anti -proliferative effect was also evaluated. Thus, two distinct geographic locations within the B. tinctoria cultivation area were selected. The contents of soil pH, organic carbon, nitrogen, phosphorus, potassium and primary and secondary active metabolites in the fruits were measured. Bioactive compounds were tentatively identified by liquid chromatography-mass spectrometry (LC-MS) analysis. Qualitative and quantitative phytochemical screening revealed that major primary and secondary metabolites were more abundant in fruits from the NH region, especially in terms of total phenols and alkaloids. The data acquired highlight that soil physicochemical properties have a highly positive correlation with secondary metabolites content and bioactivity. In particular, NH ethyl acetate (NH-EAF) extracts showed higher antioxidant and anti-inflammatory activiy, as well as anti-bacterial activity against Staphylococcus aureus and Enterococcus faecium. Furthermore, NH-EAF extract showed cytotoxic effects with IC50 equal to 102.85 mu g/mL and 123.39 mu g/mL towards Raw 264.7 and Caco-2 cell lines, respectively. The results of this study confirmed that the fruit variability of phytochemicals and biological activities is due to geographical variation and ecological factors.

This study investigated the anticancer phytocompounds in leaf extracts of Oxalis latifolia Kunth. Quantitative analysis of the phytochemical composition showed high levels of primary metabolites: carbohydrates (45.11 ± 2.15 GLE mg/100 mg), proteins (28.13 ± 0.94 BSA mg/100 mg), and amino acids (13.25 ± 1.16 LE mg/100 mg). Ethyl acetate extracts had the highest concentrations of secondary metabolites, including phenolic content (122.52 ± 4.27 GAE mg/100 mg), total flavonoids (91.86 ± 2.65 QE mg/100 mg), and alkaloids (82.18 ± 0.72 COLE mg/100 mg). In addition, strong antioxidant activities were observed in the DPPH• scavenging assay (IC50 11.51 ± 2.28 µg/mL), ABTS·+ radical cation scavenging activity (97.42 ± 7.19 µM TE/g), and FRAP assay (14.34 ± 1.24 mM Fe(II)/mg). Based on preliminary analysis, the ethyl acetate extract was fractionated using thin-layer chromatography (TLC), yielding two distinct fractions with Rf values of 0.31 and 0.76. GC–MS analysis of these fractions identified 33 bioactive compounds. These fractions exhibited anticancer activity against the A549 lung cancer cell line, with IC50 values of 47.25 µg/mL and 48.31 µg/mL, as determined by the MTT assay. Furthermore, absorption, distribution, metabolism, excretion, and toxicity (ADMET) studies on 25 selected compounds indicated favorable pharmacokinetic properties and drug-likeness. In silico molecular docking showed strong binding affinities of these bioactive compounds to the p21 protein, comparable to the synthetic drug Cisplatin–quercetin. The results highlight the potential of O. latifolia in anticancer therapy, particularly through modulation of the p21 pathway, supported by in vitro cytotoxicity assessments, molecular docking, and ADMET analysis.

In recent years, functional foods have gained attention for their potential health benefits beyond basic nutrition, serving as rich sources of proteins, carbohydrates, vitamins, and dietary fiber. Enriched with bioactive compounds such as polyphenols, tannins, flavonoids, and alkaloids, these foods have shown promise in inhibiting cell signaling pathways related to proliferation, communication, and apoptosis. Key components in functional foods offer health benefits. Omega-3 fatty acids scavenge free radicals, reducing the risk of chronic diseases like heart disease and cancer. Antioxidants in foods such as sesame seed oil promote cardiovascular health and have anti-inflammatory effects. Curcumin inhibits cancer by suppressing inflammatory cytokines. Understanding their mechanisms of action is crucial for realizing therapeutic benefits. However, identifying suitable bioactive substances and comprehending their role in disease prevention are challenging. As we enter the functional food era, this review aims to present the current state of functional foods and nutraceuticals, exploring their applications for human wellness. Recognizing nutrition's individualized nature of nutrition is a central theme, emphasizing the importance of tailored dietary recommendations based on diverse responses to these functional elements, ensuring that personalized approaches are aligned with individual health goals.

Food preparation involves the blending of various food ingredients to make more convenient processed food products. It is a long chain process, where each stage posing a risk of accumulating hazardous contaminants in these food systems. Protecting the public health from contaminated foods has become a demanding task in ensuring food safety. This review focused on the causes, types, and health risks of contaminants or hazardous chemicals during food processing. The impact of cooking such as frying, grilling, roasting, and baking, which may lead to the formation of hazardous by-products, including polycyclic aromatic hydrocarbons (PAHs), heterocyclic amines (HCAs), acrylamide, advanced glycation end products (AGEs), furan, acrolein, nitrosamines, 5-hydroxymethylfurfural (HMF) and trans-fatty acids (TFAs). Potential health risks such as carcinogenicity, genotoxicity, neurotoxicity, and cardiovascular effects are emerging as a major problem in the modern lifestyle era due to the increased uptakes of contaminants. Effects of curing, smoking, and fermentation of the meat products led to affect the sensory and nutritional characteristics of meat products. Selecting appropriate cooking methods include temperature, time and the consumption of the food are major key factors that should be considered to avoid the excess level intake of hazardous contaminants. Overall, this study underscores the importance of understanding the risks associated with food preparation methods, strategies for minimizing the formation of harmful compounds during food processing and highlights the need for healthy dietary choices to mitigate potential health hazards.

Legumes have played an essential part as food and feed, but their productivity is inadequate to deliver the protein needs of a rapidly growing human population and livestock sectors. In numerous places around the world, legumes are the peculiar source of protein which essential for the human consumption. They are typically used as a supplement to other dietary proteins. As legumes have the richest protein source, the health experts and consumers demand increased rapidly. Simultaneously, the inhabitants of humans in the universe increased promptly. Meanwhile, fluctuation in biotic and also abiotic components is an extensive constraint for crop growth. It may provoke stress which results in the reduction of yield. However, the legume shows recalcitrant nature, these are the major root cause of the shortage of food. The biotechnology provides a solution to rectify all those issues and genetic transformation plays a consequential impact on enhancement of the legume productivity. This review clearly emphasizes the constraints of legume productivity, the strategy used, skill requirement and current advances in biotechnology for legume improvement.

Two lipid sensors, CD36 and GPR120, are crucial for the orosensory detection of fat taste and for mediating fat preference. However, the mechanism by which endogenous lipid (FAHFA) binds to CD36 to initiate intracellular signaling remains unexplained. Hence, the primary objective of this study is to investigate the binding mechanism of FAHFA to CD36 and its role in isolated mouse taste bud cells (mTBCs). The Schrodinger platform was used to assess the molecular dynamics of protein and ligand interactions, and an in vitro experiment was used to validate the findings. Based on the docking score of the ligand, the molecular mechanistic activities of the targeted complexes, CD36-5-POHSA (-8.2 kcal/mol), were investigated using the dynamic simulation. In comparison to linoleic acid (LA), POHSA rapidly increased [Ca2+]i via acting on CD36, and 5-POHSA treatment in mTBCs activated src-kinase at 20 μM. CD36 siRNA transfection in TBCs downregulate the CD36 protein expression as well as [Ca2+]i flux. This study suggests that 5-POHSA may help combat taste abnormalities and the adverse effects of obesity by binding to the lingual CD36 receptor and activating the tongue-brain axis.