Plasmids for Optimizing Expression of Recombinant Proteins in <em>E. coli</em>

Plasmids are important vectors for the transfer of genetic material among microbes. The transfer of plasmids causes transmission of genes involved in pathogenesis and survival, to the host bacteria leading to their evolution and adaptation to diverse environmental conditions. A large number of plasmids of varying sizes have been discovered and isolated from various microorganisms. Plasmids are also valuable tools to genetically manipulate microbes for various purposes including production of recombinant proteins. Escherichia coli is the most preferred microbe for production of recombinant proteins, due to rapid growth rate, cost-effectiveness, high yield of the recombinant proteins and easy scale-up process. Several plasmids have been designed to optimize the expression of heterologous proteins in E. coli. In order to circumvent the issues of protein refolding, the codon usage in E. coli, the absence of post-translational modifications, such as glycosylation and low recovery of functionally active recombinant proteins, various plasmids have been designed and constructed. This chapter summarizes the recent technological advancements that have extended the use of the E. coli expression system to produce more complex proteins, including glycosylated recombinant proteins and therapeutic antibodies

Molecular identification of goat"s udder microbes and nutritional value of milk using dielectric constants

This study aims to determine which bacteria are present in goat"s udder by using gram dye, nuclear nucleic acid technology, direct 16S rRNA gene sequencing, and dielectric constants. The 16S rRNA gene is frequently sequenced in modern approaches to identifying microorganisms and molecules. Bacteria like Cohnella suwonensis, Cohnella yongneupensis, and Cohnella faecalis were isolated and Staphylococcus species like succinus, lentus, xylosus, equorum, stepanovicii, fleurettii, sciuri, kocurii, and faecalis were recorded. The constants such as dielectric constant (ε′), spectra (ε″) and dielectric loss factor (ε′+ε″) spectra, and Total Bacterial Count (TBC) from raw goat milk samples were determined employing network analyzer and plate count methods, respectively. A poor relationship was found between TBC in logarithm and permittivities at single and multiple frequencies. Linear models like multiple linear regression, ridge regression, and absolute shrinking were used to determine TBC based on effective dielectric constant and spectra and their combination; and an excellent TBC performance was determined. In conclusion, various microbes found and isolated were, Staphylococcus succinus, S. lentus, S. xylosus, S. equorum, S. stepanovicii, S. fleurettii, S. sciuri, S. kocurii, Cohnella suwonensis, C. yongneupensis, and C. faecalis. This study emphasizes the importance of improved hygienic procedures in ensuring food safety by emphasizing the importance of avoiding contamination and using the appropriate types of processing equipment

Urban Quality of Life in Nasr City: An Assessment of the Reformation Project of the Built Environment

Large-scale urban renewal initiatives are undertaken by governments in cities under pressure to improve residents' quality of life (QoL) and mitigate economic and environmental issues. In 2019, as a part of the Egypt 2030 vision, the government initiated a road network reformation project to solve mobility issues in the districts linking the east and west of Cairo, including Nasr City District. To investigate if the built environment of Nasr City District is still resident-friendly, this research is being undertaken to examine residents' satisfaction with the reformation project, which had an impact on their QoL

A Sensitive and Rapid Assay for Investigating Vertical Transmission of Hepatitis B Virus via Male Germ Line Using EGFP Vector as Reporter

Hepatitis B virus (HBV) constitutes a serious menace to man. DNA recombination and sequencing, interspecific in vitro fertilization, single-embryo PCR and RT-PCR were employed to establish a sensitive and rapid assay for exploring the vertical transmission of viruses via male germ line. Plasmid pIRES2-EGFP-HBs which expressed enhanced green fluorescent protein as reporter for the expression of hepatitis B virus S gene was successfully constructed and confirmed by PCR, EcoR I and Sal I digestion, and DNA sequencing. After exposure to the plasmid, human spermatozoa were used to fertilize with zona-free hamster ova. Two-cell embryos were collected and classified into group A with green fluorescence and group B without green fluorescence under fluorescence microscope. The results showed that HBs DNA positive bands were detected in the embryos with green fluorescence (PCR and RT-PCR) and positive control (PCR) indicating expression of pIRES2-EGFP-HBs, and not observed in the embryos without green fluorescence and negative controls (PCR and RT-PCR) indicating no pIRES2-EGFP-HBs in the cells. The advantages and application foreground of this assay for study on vertical transmission of viruses such as HCV, HIV, HPV, and SARS via germ line were discussed

Molecular identification of microbial contaminations in the fitness center in Makkah region

As the condition increases and seeks to remain healthy, the number of people who plan to join a fitness center or "gym" has increased markedly. From where this individual does understand, the study of the variety of bacteria showing the stolen has led him to care for people, with a popular fitness center located in the province of Makka, Saudi Arabia. Different bacteria must be eliminated from other gyms in Makkah, Saudi Arabia, in a total of three areas 46 two sports equipment. Both types and characteristics of bacteria, while some have been tested in hemolytic surgery for antibiotic resistance. Corynebacterium antibiotics in different forms did not react the same; however, isolates tested for M17 and N12 showed the greatest resistance to antibiotics. Furthermore, sixteen bacterial strains of human blood β-agar displayed high hemolytic activity. In the gym isolates 2 (9 strains) followed in gym 1 (7 modes), B row hemolytic activity was highest. It is important to note that gram-positive bacteria were positive in all kinds of ways, and catalase was positive. Six strains belonging to the genus Bacillus, Brachybacterium, Geobacillus, Microbacterium, Micrococcus, and Staphylococcus and other pathogenic bacteria were known as possible individuals to use the morphological, biochemical, and rRNA gene of the 16S series. In general, this research illustrates the health and fitness centers in the individual being studied and the risks that are considered necessary to periodically study possible microbial contamination in the mixture in the gym to ensure people's protection

Role of static fluid MR urography in detecting post urinary diversion complications

Aim of work: The aim of the study was to assess the diagnostic performance of static MR urography in detection of post cystectomy complications &amp; the ability of static fluid MR urography in visualization of urinary tract segments.Material &amp; methods: We prospectively reviewed 21 MR urograms with urinary diversion. The most common surgical procedures included Ileal conduit &amp; Ileocecal neobladder diversion.Material &amp; methods: Magnetic resonance urography examinations were performed with 1.5-T MR scanners. T2 weighted (static fluid) MR urography techniques were done, in addition to conventional T1- and T2-weighted axial and coronal sequences. Urinary tract was divided in different parts: pelvicalyceal systems, upper, mid and lower ureteric segments &amp; the reservoir or conduit Imaging features of the urinary collecting systems were evaluated for their visualization and complications detection.Results: T2-weighted MR urography could demonstrate 95% of urinary tract segments &amp; together with conventional MR sequences all urinary tract segments can be visualized. Urinary diversion related complications were encountered included in 15 patients (71.4%) &amp; no urological complications were seen in 6 patients (28.6%).Conclusion: Comprehensive T2-weighted MR urography is an effective imaging method for the visualization of the urinary system and detection of early and late postoperative complications in patients with urinary diversion.Keywords: MR urography, Urinary diversion, Cancer bladde

Assessing Drought Tolerance of Newly Developed Tissue-Cultured Canola Genotypes under Varying Irrigation Regimes

This article belongs to the Special Issue Crop Tolerance under Biotic and Abiotic Stresses[Abstract] Drought is a major abiotic stress that greatly affects canola growth, production, and quality. Moreover, water scarcity is projected to be more severe and frequent as a result of climate change, in particular in arid environments. Thereupon, developing drought-tolerant and high-yielding canola genotypes has become more critical to sustaining its production and ensuring global food security with the continuing population growth. In the present study, ten canola genotypes comprising six developed tissue-cultured canola genotypes, two exotic genotypes, and two commercial cultivars were evaluated under four irrigation regimes. The applied irrigation regimes were well-watered (100% crop evapotranspiration, ETc), mild drought (80% ETc), moderate drought (60% ETc), and severe drought (40% ETc) conditions. Drought-stress treatments (80, 60, and 40% ETc) gradually reduced the chlorophyll content, relative water content, flowering time, days to maturity, plant height, number of pods, number of branches, seed yield, and oil percentage, and increased proline, phenolic, anthocyanin, and glycine betaine contents. The evaluated genotypes exhibited varied responses to drought-stress conditions. The developed tissue-cultured genotypes T2, T3, and T1, as well as exotic genotype Torpe, possessed the highest performance in all evaluated parameters and surpassed the other tested genotypes under water-deficit conditions. Overall, our findings elicited the superiority of certain newly developed tissue-cultured genotypes and exotic ones compared with commercial cultivars, which could be exploited in canola breeding under water-deficit conditions.This research was funded by the Researchers Supporting Project number (RSPD-2023R730), King Saud University, Riyadh, Saudi ArabiaKing Saud University (Riad, Arabia Saudí); RSPD-2023R73

Comparative molecular studies of halophilic bacteria from saline water and soil in the Saudi environment

Halophilic bacteria are a microorganism that grows optimally in the presence of the very high concentration of sodium chloride. Halophiles are vital sources of various enzymes including hydrolases, which are very stable and catalytically highly efficient at high salt concentration and other extreme conditions such as high temperature, pH and presence of organic solvents.  Several hydrolases such as amylases, proteases, and lipases have been obtained from halophilic bacteria and are commonly used for various industrial applications. We initiated a screening project to isolate and characterize the halophilic bacteria from the Red Sea, which is one of the saltiest bodies of water in the world. Water and soil samples, collected from the Red Sea coast, Jeddah, Saudi Arabia, were screened for isolation of halophilic bacteria. Ten bacterial isolates were obtained, which were characterized by biochemical tests and 16S rRNA gene sequencing. Hydrolase producing bacteria among the isolates were screened by plate assay on starch and gelatin agar plates for amylase and protease, respectively.  Two bacterial isolates i.e Bacillus haynesii and Enterobacter cloacae subsp. were found to possess significant amylase and protease activity. Further characterization of both the strains is in progress

Hepatobiliary manifestations following two-stages elective laparoscopic restorative proctocolectomy for patients with ulcerative colitis: A prospective observational study

BACKGROUNDHepatobiliary manifestations occur in ulcerative colitis (UC) patients. The effect of laparoscopic restorative proctocolectomy (LRP) with ileal pouch anal anastomosis (IPAA) on hepatobiliary manifestations is debated.AIMTo evaluate hepatobiliary changes after two-stages elective laparoscopic restorative proctocolectomy for patients with UC.METHODSBetween June 2013 and June 2018, 167 patients with hepatobiliary symptoms underwent two-stage elective LRP for UC in a prospective observational study. Patients with UC and having at least one hepatobiliary manifestation who underwent LRP with IPAA were included in the study. The patients were followed up for four years to assess the outcomes of hepatobiliary manifestations.RESULTSThe patients' mean age was 36 +/- 8 years, and males predominated (67.1%). The most common hepatobiliary diagnostic method was liver biopsy (85.6%), followed by Magnetic resonance cholangiopancreatography (63.5%), Antineutrophil cytoplasmic antibodies (62.5%), abdominal ultrasonography (35.9%), and Endoscopic retrograde cholangiopancreatography (6%). The most common hepatobiliary symptom was Primary sclerosing cholangitis (PSC) (62.3%), followed by fatty liver (16.8%) and gallbladder stone (10.2%). 66.4% of patients showed a stable course after surgery. Progressive or regressive courses occurred in 16.8% of each. Mortality was 6%, and recurrence or progression of symptoms required surgery for 15%. Most PSC patients (87.5%) had a stable course, and only 12.5% became worse. Two-thirds (64.3%) of fatty liver patients showed a regressive course, while one-third (35.7%) showed a stable course. Survival rates were 98.8%, 97%, 95.8%, and 94% at 12 mo, 24 mo, 36 mo, and at the end of the follow-up.CONCLUSIONIn patients with UC who had LRP, there is a positive impact on hepatobiliary disease. It caused an improvement in PSC and fatty liver disease. The most prevalent unchanged course was PSC, while the most common improvement was fatty liver disease