A Cas9-based toolkit to program gene expression in Saccharomyces cerevisiae

Despite the extensive use of Saccharomyces cerevisiae as a platform for synthetic biology, strain engineering remains slow and laborious. Here, we employ CRISPR/Cas9 technology to build a cloning-free toolkit that addresses commonly encountered obstacles in metabolic engineering, including chromosomal integration locus and promoter selection, as well as protein localization and solubility. The toolkit includes 23 Cas9-sgRNA plasmids, 37 promoters of various strengths and temporal expression profiles, and 10 protein-localization, degradation and solubility tags. We facilitated the use of these parts via a web-based tool, that automates the generation of DNA fragments for integration. Our system builds upon existing gene editing methods in the thoroughness with which the parts are standardized and characterized, the types and number of parts available and the ease with which our methodology can be used to perform genetic edits in yeast. We demonstrated the applicability of this toolkit by optimizing the expression of a challenging but industrially important enzyme, taxadiene synthase (TXS). This approach enabled us to diagnose an issue with TXS solubility, the resolution of which yielded a 25-fold improvement in taxadiene production

Agricultural Waste Valorization for Nanoparticles Synthesis and Enhancement of Vapour Compression Refrigeration System’s Performance

Waste management has been a major concern in the society and agricultural wastes can be utilized in the synthesis of nanoparticles and deployed in the vapour compression refrigeration system (VCRS) to enhance its performance. This study analysed the thermophysical properties, performance, energy consumption, pull-down time, and capacities of VCRS using bio-nanoparticles produced from orange and pineapple peels. Eco-friendly refrigerants R600a and R134a with pure polyolester (POE) as the lubricating oil for the compressor were used. The nanolubricants were dispersed in three volume fractions of 0.05%, 0.10% and 0.20% concentration in the lubricant using the two-step method. The degradation of nanolubricants were analysed by examining the thermophysical properties of the nanolubricants before and after use in the VCRS. At 0.2% volume concentration, optimum COP of 6.31 and 5.01 were obtained for pineapple and orange peels respectively for R600a. The nanolubricants of orange peels with the volume fraction of 0.2% had the best pull-down time with a temperature of-2oC. The lowest power consumption was observed for 0.1% volume concentration of pineapple nanolubricants while 0.2% volume concentration of orange nanolubricants was observed to have the least power consumption. Considering the R134a refrigerant, the volume concentration with the optimum COP was 0.1 vol% concentration for the orange bio-based nanolubricants with an increase in the COP of 36.3% when compared with pure R134a while 0.2 vol% had the best pull-down time with a temperature of-3oC. There was a 14.2% drop in the power consumption of 0.1 vol% concentration of pineapple nanolubricants when compared to the various concentrations of the bio-based nanolubricants. From this study, the optimum performance was observed at 0.20 vol% concentration for the orange and pineapple nanolubricants with a relatively less power consumption. R600a refrigerant can completely replace R134a in its use in refrigeration systems and achieve similar pull-down time and coefficient of performance when bio-nanolubricants are utilized in the systems

Spectrum of anxiety and depression reported in reproductive-aged women diagnosed with gynaecological disorders at a tertiary healthcare facility in Ghana

Background: Patients with gynaecological disorders often suffer from psychological disorders including anxiety and depression. Although depression and anxiety have been studied in Ghana, data regarding the prevalence of these disorders in patients with gynaecological disorders is non-existent. The aim of the study was to investigate the prevalence of anxiety and depression in reproductive-aged women diagnosed with gynaecological disorders.Methods: Cross-sectional observational study was conducted at the Gynaecology Clinic of Korle-Bu Teaching Hospital, a tertiary health facility in Accra, Ghana. Patients of reproductive age seeking gynaecological care at the facility from December 2018 to January 2019 were assessed for anxiety and depression using the Generalized anxiety disorder (GAD) questionnaire and the Beck depression inventory (BDI) respectively. Sociodemographic and clinical information was gathered as well.Results: Of the 120 patients interviewed (mean age 34.33±0.66), 36.7% were depressed while 51.6% were reported anxiety disorders. Patients aged 35-45 years had the highest prevalence of anxiety (24.58%) and depression (29.18%). Again, prevalence rates were highest among respondents with senior high school as the highest educational qualification, (anxiety (22.15%); depression (24.20%). Patients suffering from pelvic floor disorder recorded the highest prevalence of anxiety (11.40%) and depression (13.77%). There was a significant association between depression and gynaecological disorders [χ2(25) =53.915, p=0.001, CI=95%], but there was not enough evidence of an association between anxiety and gynaecological disorders [χ2(15) =22.791, p=0.089, CI=95%].Conclusions: Anxiety and depression are prevalent amongst women in their reproductive age diagnosed presenting with gynaecological disorders and there is a significant association between gynaecological disorders and the prevalence of depression

A review of pharmacological effects of xylopic acid

Xylopic acid (15β-acetyloxy-kaur-16-en-19-oic acid) is a kaurene diterpene that can be obtained from various Xylopia spp. Xylopic acid has demonstrated several pharmacological activities in vitro and in vivo. The compound has shown promising effect as a potent analgesic, anti-inflammatory and anti-allergic agent. Xylopic acid is a CNS depressant and was able to ameliorate anxiety-like symptoms in mice in addition to its neuroprotective effects. Deleterious effects of xylopic acid on the reproductive system of mice have been well documented but extensive toxicity study detailing effect of the acid upon chronic exposure needs to be determined. Due to the heavy consumption of X. aethiopica fruits, it is recommended that the pharmacokinetics of xylopic acid be determined to ascertain the possible food-drug interaction that may occur when conventional drugs are taken together with foods containing xylopic acid

Structural brain preservation: a potential bridge to future medical technologies

When faced with the prospect of death, some people would prefer a form of long-term preservation that may allow them to be restored to healthy life in the future, if technology ever develops to the point that this is feasible and humane. Some believe that we may have the capacity to perform this type of experimental preservation today—although it has never been proven—using contemporary methods to preserve the structure of the brain. The idea is that the morphomolecular organization of the brain encodes the information required for psychological properties such as personality and long-term memories. If these structures in the brain can be maintained intact over time, this could theoretically provide a bridge to access restorative technologies in the future. To consider this hypothesis, we first describe possible metrics that can be used to assess structural brain preservation quality. We next explore several possible methods to preserve structural information in the brain, including the traditional cryonics method of cryopreservation, as well as aldehyde-stabilized cryopreservation and fluid preservation. We focus in-depth on fluid preservation, which relies on aldehyde fixation to induce chemical gel formation in a wide set of biomolecules and appears to be a cost-effective method. We describe two theoretical recovery technologies, alongside several of the ethical and legal complexities of brain preservation, all of which will require a prudent approach. We believe contemporary structural brain preservation methods have a non-negligible chance of allowing successful restoration in the future and that this deserves serious research efforts by the scientific community

Forefoot plantar multilobular noninfiltrating angiolipoma: a case report and review of the literature

<p>Abstract</p> <p>Background</p> <p>Soft tissue tumors of the feet are uncommon and there have been very few reports of large series in the literature. These tumors continue to present the clinician with one of the most difficult problems in medicine.</p> <p>Case presentation</p> <p>We present a case of a large multilobular noninfiltrating angiolipoma at the plantar surface of the forefoot. Only three cases occurring at the foot have been previously described. We report this new case due to unusual location of the tumor, the long duration (25 years) of its existence and the unique surgical approach for the tumor excision.</p> <p>Conclusion</p> <p>Surgical excision is the treatment of choice and adjuvant radiotherapy is indicated in select cases.</p

Disruption of Growth Hormone Receptor Prevents Calorie Restriction from Improving Insulin Action and Longevity

Most mutations that delay aging and prolong lifespan in the mouse are related to somatotropic and/or insulin signaling. Calorie restriction (CR) is the only intervention that reliably increases mouse longevity. There is considerable phenotypic overlap between long-lived mutant mice and normal mice on chronic CR. Therefore, we investigated the interactive effects of CR and targeted disruption or knock out of the growth hormone receptor (GHRKO) in mice on longevity and the insulin signaling cascade. Every other day feeding corresponds to a mild (i.e. 15%) CR which increased median lifespan in normal mice but not in GHRKO mice corroborating our previous findings on the effects of moderate (30%) CR on the longevity of these animals. To determine why insulin sensitivity improves in normal but not GHRKO mice in response to 30% CR, we conducted insulin stimulation experiments after one year of CR. In normal mice, CR increased the insulin stimulated activation of the insulin signaling cascade (IR/IRS/PI3K/AKT) in liver and muscle. Livers of GHRKO mice responded to insulin by increased activation of the early steps of insulin signaling, which was dissipated by altered PI3K subunit abundance which putatively inhibited AKT activation. In the muscle of GHRKO mice, there was elevated downstream activation of the insulin signaling cascade (IRS/PI3K/AKT) in the absence of elevated IR activation. Further, we found a major reduction of inhibitory Ser phosphorylation of IRS-1 seen exclusively in GHRKO muscle which may underpin their elevated insulin sensitivity. Chronic CR failed to further modify the alterations in insulin signaling in GHRKO mice as compared to normal mice, likely explaining or contributing to the absence of CR effects on insulin sensitivity and longevity in these long-lived mice

Histone Variants and Their Post-Translational Modifications in Primary Human Fat Cells

Epigenetic changes related to human disease cannot be fully addressed by studies of cells from cultures or from other mammals. We isolated human fat cells from subcutaneous abdominal fat tissue of female subjects and extracted histones from either purified nuclei or intact cells. Direct acid extraction of whole adipocytes was more efficient, yielding about 100 µg of protein with histone content of 60% –70% from 10 mL of fat cells. Differential proteolysis of the protein extracts by trypsin or ArgC-protease followed by nanoLC/MS/MS with alternating CID/ETD peptide sequencing identified 19 histone variants. Four variants were found at the protein level for the first time; particularly HIST2H4B was identified besides the only H4 isoform earlier known to be expressed in humans. Three of the found H2A potentially organize small nucleosomes in transcriptionally active chromatin, while two H2AFY variants inactivate X chromosome in female cells. HIST1H2BA and three of the identified H1 variants had earlier been described only as oocyte or testis specific histones. H2AFX and H2AFY revealed differential and variable N-terminal processing. Out of 78 histone modifications by acetylation/trimethylation, methylation, dimethylation, phosphorylation and ubiquitination, identified from six subjects, 68 were found for the first time. Only 23 of these modifications were detected in two or more subjects, while all the others were individual specific. The direct acid extraction of adipocytes allows for personal epigenetic analyses of human fat tissue, for profiling of histone modifications related to obesity, diabetes and metabolic syndrome, as well as for selection of individual medical treatments

The epidemiology and factors associated with nocturnal enuresis among boarding and daytime school children in southeast of Turkey: a cross sectional study

<p>Abstract</p> <p>Background</p> <p>Nocturnal enuresis is an important problem among young children living in Turkey. The purpose of this study was to determine the possible differences in the prevalence of enuresis between children in boarding school and daytime school and the association of enuresis with sociodemographic factors.</p> <p>Methods</p> <p>This was a cross-sectional survey. A total of 562 self-administered questionnaires were distrubuted to parents from two different types of schools. One of them was a day-time school and the other was a boarding school. To describe enuresis the ICD-10 definition of at least one wet night per month for three consecutive months was used. Chi-square test and a logistic regression model was used to identify significant predictive factors for enuresis.</p> <p>Results</p> <p>The overall prevalence of nocturnal enuresis was 14.9%. The prevalence of nocturnal enuresis declined with age. Of the 6 year old children 33.3% still wetted their beds, while the ratio was 2.6% for 15 years-olds. There was no significant difference in prevalence of nocturnal enuresis between boys and girls (14.3% versus 16. 8%). Enuresis was reported as 18.5% among children attending day time school and among those 11.5% attending boarding school (p < 0.05). Prevalence of enuresis was increased in children living in villages, with low income and with positive family history (p < 0.05). After multivariate analysis, history of urinary tract infection (OR = 2.02), age (OR = 1.28), low monthly income (OR = 2.86) and family history of enuresis (OR = 3.64) were factors associated with enuresis. 46.4% of parents and 57.1% of enuretic children were significantly concerned about the impact of enuresis.</p> <p>Conclusion</p> <p>Enuresis was more frequent among children attending daytime school when compared to boarding school. Our findings suggest that nocturnal enuresis is a common problem among school children, especially with low income, smaller age, family history of enuresis and history of urinary tract infection. Enuresis is a pediatric public health problem and efforts at all levels should be made such as preventive, etiological and curative.</p