A review on molecular scissoring with CRISPR/Cas9 genome editing technology.

Genome editing is a technology to make specific changes in the DNA of a cell or an organism. It has significantly altered the landscape of life sciences, facilitating the establishment of exceedingly customized genetic modifications. Among various genome editing technologies, the CRISPR/Cas9 system, a specific endonuclease induces a double stranded DNA break and enabling modifications to the genome, has surfaced as a formidable and adaptable instrument. Its significance cannot be overstated, as it not only allows for the manipulation of genomes in model organisms but also holds great potential for revolutionary advances in medicine, particularly in treating genetic diseases. This review paper explores the remarkable journey of CRISPR/Cas9, its natural function, mechanisms, and transformative impact on genome editing and finally the use of artificial intelligence and other intelligent manufacturing tools used. The introduction provides the background on genome editing, emphasizing the emergence and significance of CRISPR/Cas9. Subsequent sections comprehensively elucidate its natural function, disease modeling, agriculture, and biotechnology, address therapeutic applications, and ongoing clinical trials while also discussing prospects and ethical implications. We summarized the key findings, indicating that CRISPR/Cas9 has empowered the creation of disease-specific animal models. This provides invaluable insights into pathogenic mechanisms and opens new avenues for drug discovery, reaffirming the transformative impact of CRISPR/Cas9 on genome editing. Finally we discussed the importance of continued research and collaboration for comprehensive utilization of the inherent capabilities of this molecular precision tool in shaping forthcoming advancements.

Mitigating NaCl stress in Vigna radiata L. cultivars using Bacillus pseudomycoides.

Salt stress is one of the significant abiotic stress factors that exert harmful effects on plant growth and yield. In this study, five cultivars of mung bean (Vigna radiata L.) were treated with different concentrations of NaCl and also inoculated with a salt-tolerant bacterial strain to assess their growth and yield. The bacterial strain was isolated from the saline soil of Sahiwal District, Punjab, Pakistan and identified as Bacillus pseudomycoides. Plant growth was monitored at 15-days interval and finally harvested after 120 days at seed set. Both sodium and potassium uptake in above and below-ground parts were assessed using a flame photometer. Fresh and dry mass, number of pods, seeds per plant, weight of seeds per plant and weight of 100 seeds reduced significantly as the concentration of NaCl increased from 3 to 15 dSm-1. There was a significant reduction in the growth and yield of plants exposed to NaCl stress without bacterial inoculum compared to the plants with bacterial inoculum. The latter plants showed a significant increase in the studied parameters. It was found that the cultivar Inqelab mung showed the least reduction in growth and yield traits among the studied cultivars, while Ramzan mung showed the maximum reduction. Among all the cultivars, maximum Na+ uptake occurred in roots, while the least uptake was observed in seeds. The study concludes that NaCl stress significantly reduces the growth and yield of mung bean cultivars, but Bacillus pseudomycoides inoculum alleviates salt stress. These findings will be helpful to cultivate the selected cultivars in soils with varying concentrations of NaCl.

Tamarindus indica seed polysaccharide-copper nanocomposite: An innovative solution for green environment and antimicrobial studies.

The purpose of this study was to synthesize ecofriendly nano-composite in which agricultural waste (seeds of Tamarindus indica) was used to synthesize tamarind seed polysaccharides (TSP) and its composite with copper nanoparticles (Cu-NPs) for the purpose of green and clean environment as well as reduction of green-house gases. Confirmation of extracted TSP, synthesized nanocomposite was carried out using FTIR, SEM, PXRD and EDX techniques. In FTIR analysis TSP gives a strong broad peak at 3331 cm-1 due to -OH group and in case of composite its intensity is reduced which might be due to the interactions between -OH and Cu+2 ions. SEM analysis gives that TSP have irregular and rough surface while Cu-NPs exhibited spherical morphology and composite showed clustering of spherical shape to rough surface. EDX analysis quantitatively represented copper having atomic ratio 0.57 % which confirms the synthesis of composite. Furthermore, synthesized composite demonstrated excellent antibacterial activity against gram-positive (S.aureus) and gram-negative bacteria (E.coli) even greater than standard medicine (ciprofloxacin). From this study it was revealed that agriculture waste can be utilized to make environment green as well as synthesized composite from agricultural waste seed also displayed excellent antimicrobial activities which directs that they can be utilized in medical field. This study aims to assess the antimicrobial properties of the nanocomposite, aiming to contribute to the development of effective antimicrobial agents. Through these objectives, the research seeks to bridge the gap between green technology and antimicrobial efficacy, offering a promising avenue for both environmental conservation and healthcare advancements.

The Global Impact of COVID-19: A Comprehensive Analysis of Its Effects on Various Aspects of Life.

In this study COVID-19 effects on different aspects of life that how this virus created a mess in every discipline of life starting from a small tuck shop of a street to a huge business with a chain between different countries; and some preventive measures are also suggested. Not only mental healthiness as well as physical health of people was also disturbed to a large extent. People being quarantined did not do any practice and had nothing to do, their boredom made them mentally and physically inactive. For minimization the effect of this pandemic on mental healthiness, interventions were practiced and psychological support systems were developed to help mentally effected people; on the other hand, to improve physical health the hospital workers worked day and night in return they got affected too either mentally or physically. Many of the youngsters started alcohol consumption during quarantine. Because of the closure of educational institutes, the students were sent back to their homes where there was no proper guidance for them and they lost their interests in studies; and in a sense educational impact of COVID-19 was also unbearable. Agricultural system was affected badly and the whole world passed through a huge economic loss. The flights and traffic were blocked throughout the world, and it is the only positive impact that COVID-19 led to the environment by improving water and air quality as there was a remarkable reduction in the emission of greenhouse gases.

Novel approach to water-efficient bulk industrial textile printing production of cotton fabric.

Water scarcity poses a global threat in climate change era, and regrettably, the textile processing industry is squandering a significant volume of water during bulk production. This research focused on a sustainable water-saving approach in the printing of cotton fabric by modifying the reactive printing recipe and methodology. Three modified recipes (X, Y, Z) and one controlled recipe (C) were tested using reactive dyes. The conventional reactive printing recipe (Control) includes sodium alginate, urea, mild oxidizing agent, and sodium bicarbonate. In contrast, the modified recipe trials incorporated an acrylic-based synthetic thickener in the replacement of sodium alginate (alone and in combination with sodium alginate). A total of four recipes (one controlled conventional and three modified recipes) were examined using three reactive dyes at two dose levels (2 % and 4 %). Various characterization techniques, including shade variation, color penetration into the fabric, sharpness of the edges, color tinting on the adjacent white fabric, perspiration fastness (both acidic and alkaline), washing fastness, rubbing fastness, and fabric hardness, affirmed that Y recipe yielded the best results in fabric testing, cost reduction, and water conservation. This research represents a pioneering contribution to the printing industry with novel recipes.

Green synthesis of Eucalyptus globulus zinc nanoparticles and its use in antimicrobial insect repellent paint formulation in bulk industrial production.

Mitigating climate change can be achieved by opting for sustainable, plant-based materials instead of relying on hazardous chemicals that come with various side effects. Various natural plant extracts find widespread application in synthesizing insect-repellent coatings, particularly in industries such as paint manufacturing. The increasing demand for these coatings has led us to find out the effects of different plant extracts for the efficient preparation of paints with more advanced impacts and low cost. For this purpose, zinc nanoparticles of Eucalyptus globulus L. and its extracts were used in this study due to their remarkable biocidal and antimicrobial activities. The extract was prepared by the process of oven-drying and heating followed by their filtration. Then, they were subjected to different phytochemical tests that were performed in which plant material did not contain flavonoids and glycol. The comparison of the size of nanoparticles was visible during the weighing which was found to be 4.451 mg. Advanced characterization techniques like FTIR, UV visible spectroscopy, and particle size analysis were adopted for the analysis of nanoparticles of plant extract. The FTIR analysis of the plant material was reported to lie in the range of 1000-1800 cm-1. On the other hand, the results of UV visible spectroscopy of nanoparticles of plant extract showed absorption peaks around 300 nm. The produced material was integrated into paint formulations to impart insect-repellent and antibacterial characteristics. Painted panels exhibited notable antibacterial efficacy, presenting an inhibition zone of 0.7 cm for Escherichia coli and 0.3 cm for Staphylococcus aureus when utilizing biocide. Plant nanoparticles yielded inhibition zones of 1 and 1.2, while aqueous extract resulted in zones of 0.2 and 0.5, respectively. A thorough evaluation of the paint's color attributes, including ΔL, Δa, Δb, and ΔE, indicated noteworthy differences. The CMC ΔE values from the trials exceeded 1, indicating a substantial change in shade. The batches of paints containing E. globulus extracts and nanoparticles were found to be lighter in color specifically green and yellow colors. Their antimicrobial and insect repellant activity was tested using the mosquitos of Aedes aegypti with an age of 4-5 weeks, revealing that formulations with plant extracts exhibited a 61 % effective period, greater than the 7 % observed in non-biocidal formulations. The paint responded best towards these mosquitoes in terms of repellency and the ultimate target of this study was achieved.

Bulk industrial production of sustainable cellulosic printing fabric using agricultural waste to reduce the impact of climate change.

In this research paper, a novel process was developed for reactive printing of cotton fabric, with the objective of producing a high-quality printed fabric that is sustainable, eco-friendly, and low-cost which will ultimately reduce the impact of climate change. The study incorporated substituted tamarind polysaccharide (STP) obtained from agricultural waste, trichloro-ethanoic acid (TCEA), and polyethylene glycol (PEG-400) in the reactive printing paste. Tamarind starch was extracted from the seeds having 72 % yield, and substitution was performed to use it as a thickener in the printing paste. The conventional printing system was formulated with sodium alginate, urea, and sodium bicarbonate at dose levels of 2 %, 15 %, and 2.5 %, respectively, while the modified recipe was formulated with STP and TCEA at 5 % and 3 % dose levels, respectively along with varying doses of PEG-400 (0 %, 1 %, and 2 %) in novel prints. Various factors such as shade comparison, penetration, staining on the white ground, washing, rubbing, light and perspiration fastness, sharpness of edges, and fabric hardness were evaluated for all the recipes. The study demonstrated that the optimal outcomes were obtained with a 2 % PEG-400 dose level. This study represents a significant contribution to sustainable textile production, as tamarind agriculture waste was used as a raw material, which is an environmentally friendly alternative of sodium alginate that reduces the wastewater load. Additionally, PEG-400 was utilized as a nitrogen-free solubilizing moisture management substitution of urea for printing, while TCEA dissociated at high temperature to make alkaline pH during curing of the printed fabric to replace sodium bicarbonate. This research is a novel contribution to the printing industry, as these three constituents have not been previously used together other than this research group, in the history of reactive printing.

Application of Synergistic Phenomena for Enhanced Production of Alpha 1,4 D- Glucan Glucanohydrolase Using Submerged Fermentation.

The study revealed enhanced production of α 1, 4 D glucan glucanohydrolase utilizing the synergistic phenomena of bacterial hetero-culture. For this purpose, 101 hetero-cultures were screened qualitatively and quantitatively. The bacterial hetero-culture showing the highest amylolytic potential was identified as Bacillus subtilis and Bacillus amyloliquefeciens by 16S rDNA sequencing technique. Different fermentation media were evaluated and M 5 gave maximum GGH production. Different physicochemical parameters like incubation time, temperature, initial pH and inoculum size was optimized. The optimal enzyme production was obtained at 24 h, 37oC, pH 7.0 and 3% inoculum size. Glucose (3%), ammonium sulphate (1.5%) and yeast extract (2.0%) was selected as best carbon and nitrogen source, respectively. The novelty of the present piece of research was the application of the hetero-culture technique for enhanced GGH production using submerged fermentation which was not experienced before with these strains.

Process optimization for enhanced production of cellulases form locally isolated fungal strain by submerged fermentation

Cellulase has myriad applications in various sectors like pharmaceuticals, textile, detergents, animal feed and bioethanol production, etc. The current study focuses on the isolation, screening and optimization of fungal strain through one factor at a time technique for enhanced cellulase production. In current study sixteen different fungal cultures were isolated and the culture which quantitatively exhibits higher titers of cellulase activity was identified both morphologically and molecularly by 18S rDNA and designated as Aspergillus niger ABT11. Different parameters like fermentation medium, volume, temperature, pH and nutritional components were optimized. The highest CMCase and FPase activities was achieved in 100ml of M5 medium in the presence of 1% lactose and sodium nitrate at 30 oC, pH5 after 72 hours. The result revealed A. niger can be a potential candidate for scale up studies.

Light microscopy and scanning electron microscopy of bacterial colonies isolated from ras malai samples from different markets of Lahore, Pakistan.

The current study investigates the total bacterial contamination in various packed and unpacked ras malai samples of 14 different localities of Lahore, Pakistan. The bacterial colonies such as Bacillus sp. and Gamella sp. were isolated from ras malai samples and grown on agar-broth media under sterile environmental conditions. Serial dilution technique was used to compose the replicates to get a viable count of bacteria in the samples. Results indicated that in case of packed ras malai samples, maximum bacterial count was observed in Sample 1 (422 × 10-2 to 402 × 10-6 ) and minimum bacterial count was in Sample 4 (21 × 10-2 to 9.3 × 10-6 ). For unpacked ras malai samples, maximum bacterial count was in Sample 3 (200.3 × 10-2 to 181.3 × 10-6 ) and minimum bacterial count was observed in Sample 1 (110 × 10-2 to 90.4 × 10-6 ). It was concluded that the marketed samples contain more bacterial count as compared to the standard sterilization values. Such products could possibly become the cause of many health problems in children.

Enhanced production of ß- glucosidase by locally isolated fungal strain employing submerged fermentation / Aumento da produção de ß-glucosidase por estirpe fúngica isolada localmente, por fermentação submersa

ß-glucosidase has wide spectrum of biotechnological applications in different industries including food, textile, laundry detergents, pulp and paper, pharmaceutical and biofuel industry. The present investigation related to isolation, screening, and process optimization of fungal strain for enhanced production of ß-glucosidase (BGL). For this purpose, different fungal stains were isolated from different sources including soil, fruits, bark of tree as well as from the compost. The screening of fungal strain for BGL production was carried out via submerged fermentation. All the tested strains were identified on the basis of micro and macroscopic features. The fungal strain having greater ability for BGL synthesis among tested ones wasidentified as Aspergillus niger and given the code SBT-15. The process parameter including fermentation media, temperature, pH, rate of fermentation, carbon and nitrogen sources, volume of media were optimized. Five different fermentation media were evaluatedM3medium gave maximum production. The optimal conditions for BGL production was 72 hours of incubation at 40°C, pH 6 and 50 ml fermentation medium. Glucose (1%) and ammonium sulphate(3%) were optimized as best carbon and nitrogen sources, respectively.

A ß-glicosidase possui amplo espectro de aplicações biotecnológicas em diferentes indústrias, incluindo alimentos, têxteis, detergentes para lavanderia, papel e celulose, indústria farmacêutica e de biocombustíveis, etc. A presente investigação relaciona-se ao isolamento e triagem e otimização de processos de cepas fúngicas para produção aumentada de ß- glucosidase (BGL). Para este efeito, diferentes manchas fúngicas foram isoladas a partir de diferentes fontes, incluindo solo, frutos, casca de árvore, bem como a partir do composto. A triagem da linhagem fúngica para produção de BGL foi realizada via fermentaçãosubmersa. Todas as cepas testadas foram identificadas com base em características micro e macroscópicas. A linhagem fúngica com maior capacidade de síntese de BGL entre os testados foi identificada como Aspergillus niger e recebeu o código SBT-15. O parâmetro do processo, incluindo meios de fermentação, temperatura, pH, taxa de fermentação, fontes de carbono e nitrogênio, volume de mídia foram otimizados. Cinco meios de fermentação diferentes foram avaliados. O meio M3 deu a produção máxima. As condições ótimas para a produção de BGL foram 72 horas de incubação a 40 ° C, pH 6 e 50ml de meio de fermentação. Glicose (1%) e sulfato de amônio (3%) foram otimizados como melhores fontes de carbono e nitrogênio, respectivamente.

Process optimization for pectinase production by locally isolated fungal strain using submerged fermentation / Otimização de processos para produção de pectinase por cepa fúngica localmente isolada usando fermentação submersa

The present study deals with the isolation screening and optimization of fungal strain for pectinase production. The fungal strains were isolated from different sources, including soil, fruits etc. Qualitative screening was performed on the basis of the pectin hydrolysis zone. While, quantitative screening was carried out employing submerged fermentation. Among all the strains the strains showing highest pectinolytic potential were selected identified and assigned the code Aspergillus niger ABT-5.The influence of different fermentation media on pectinase production was evaluated. The M5 medium containing 10g wheat bran, nutrient medium containing (g/l) of (NH4)2SO4 6.0, K2HPO4 6.0, KH2PO4 6.0, MgSO4.7H2O 0.1 gave the highest pectinase production. The other important physico chemical parameters including incubation period, temperature, and volume of media, size of inoculum, carbon and nitrogen sources were also optimized for pectinase production. The highest pectinase production (15.5U/ml) was obtained at 72h of incubation, pH 6, temperature 30°C, volume of media 50ml. Fructose and urea were designated as best carbon and nitrogen sources subsequently.

O presente estudo trata da triagem de isolamento e otimização da cepa fúngica para produção de pectinase. As cepas fúngicas foram isoladas de diferentes fontes, incluindo solo, frutas, etc. A triagem qualitativa foi realizada com base na zona de hidrólise da pectina. Enquanto, a triagem quantitativa foi realizada utilizando fermentação submersa. Entre todas as cepas, as cepas que apresentaram maior potencial pectinolítico foram selecionadas e atribuídas ao código Aspergillus niger ABT-5. Avaliou-se a influência de diferentes meios de fermentação na produção de pectinase. O meio M5 contendo 10g de farelo de trigo, meio nutriente contendo (g / l) de (NH4)2SO4 6.0, K2HPO4 6.0, KH2PO4 6.0, MgSO4.7H2O 0.1, proporcionou a maior produção de pectinase. Os outros parâmetros físico-químicos importantes, incluindo período de incubação, temperatura e volume dos meios, tamanho do inóculo, fontes de carbono e nitrogênio também foram otimizados para a produção de pectinase. A maior produção de pectinase (15,5U / ml) foi obtida às 72h de incubação, pH 6, temperatura 30 ºC, volume dos meios 50ml. A frutose e a ureia foram designadas como melhores fontes de carbono e nitrogênio posteriormente.

Influence of medium composition and physical factors on enhanced production of endoglucanase by locally isolated fungal strain in solid state fermentation.

Endoglucanase is one of the most important enzymes of the cellulase group.  Endoglucanase are involved in the catalytic hydrolysis of cellulose and plays a pivotal role in different sectors like pharmaceutical, textile, detergent, and food processing as well as paper and pulp industry. With consumers getting more and more aware of environmental issues, industries find enzymes as a better option over other chemical catalysts. In the current research different thermophilic fungal strains were isolated from the different sources. Qualitative screening was carried out on the basis of cellulose hydrolysis zone. The quantitative screening was carried out employing solid state fermentation.  The fungal culture, showing highest EG potential was selected identified and assigned the code Aspergillus fumigatus BBT2. Different fermentation media were evaluated and M 2 containing wheat bran gave maximum EG production. The maximal enzyme productivity was recorded in 72 hours, 40°C, pH 5, inoculum size 1.5ml, and moisture content (1:1). Glucose (1%) and peptone (1%) were optimized as best carbon and nitrogen sources, respectively.

Production of rabbit antibodies against purified Glucose oxidase

Glucose oxidase is an active oxygen species generating enzyme produced from Aspergillus niger grown in submerged fermentation. Disintegration of the mycelium resulted in high glucose oxidase activity that was subjected to ammonium sulfate precipitation at 60-85% saturation rates that resulted to 6.14 U mg -1 specific activity. Purification of enzyme by anion exchange column (DEAE-Cellulose) resulted into 22.53 U mg-1 specific activity and 10.27 fold purification. This was applied to sephadex G-200 column for gel filtration chromatography. It was observed that enzyme achieved 59.37 U mg-1of specific activity with 27.08 fold purity and 64.36% recovery. Purified glucose oxidase was injected into rabbits through intravenous route, to raise the glucose oxidase antibodies. After 30 days incubation period, the rabbits were slaughtered and serum was separated from blood. The antibodies were isolated by ammonium sulfate precipitation and confirmed by agar gel precipitation test. This could be a convenient and low cost alternate assay for the estimation of glucose oxidase in biological fluids. Moreover, such antibodies against the said enzyme could be used in various therapeutic and diagnostic applications.

Production of extracellular lipases by Rhizopus oligosporus in a stirred fermentor

The present investigation deals with the kinetics of submerged extracellular lipases fermentation by both wild and mutant strains of Rhizopus oligosporus var. microsporus in a laboratory scale stirred fermentor. Other parameters studied were inoculum size, pH, agitation and rate of aeration. It was found that the growth and lipases production was increased gradually and reached its maximum 9.07± 0.42ª U mL-1 (W) and 42.49 ± 3.91ª U mL-1 (M) after 30h of fermentation for both wild and mutant strain. There is overall increase of 109 percent (W) and 124 percent (M) in the production of extracellular lipases as compared to shake flask. Another significant finding of the present study is that the fermentation period is reduced to 30 h in case of wild and 23 h in case of mutant from 48 h in shake flask studies. The specific productivity of mutant strain (qp = 377.3 U/g cells/h) was several folds higher than wild strain. The specific production rate and growth coefficient revealed the hyperproducibility of extracellular lipases using mutant IIB-63NTG-7.

Production of extracellular lipases by Rhizopus oligosporus in a stirred fermentor.

The present investigation deals with the kinetics of submerged extracellular lipases fermentation by both wild and mutant strains of Rhizopus oligosporus var.microsporus in a laboratory scale stirred fermentor. Other parameters studied were inoculum size, pH, agitation and rate of aeration. It was found that the growth and lipases production was increased gradually and reached its maximum 9.07± 0.42(a) U mL(-1) (W) and 42.49 ± 3.91(a) U mL(-1) (M) after 30h of fermentation for both wild and mutant strain. There is overall increase of 109% (W) and 124% (M) in the production of extracellular lipases as compared to shake flask. Another significant finding of the present study is that the fermentation period is reduced to 30 h in case of wild and 23 h in case of mutant from 48 h in shake flask studies. The specific productivity of mutant strain (qp = 377.3 U/g cells/h) was several folds higher than wild strain. The specific production rate and growth coefficient revealed the hyperproducibility of extracellular lipases using mutant IIB-63NTG-7.