Browsing by Subject "qPCR"

Now showing 1 - 3 of 3
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    Detection and Quantification of Taste and Odor Producing Bacteria in Eagle Creek Reservoir
    (2019-08) Koltsidou, Ioanna; Picard, Christine J.; Druschel, Gregory K.; Anderson, Gregory G.
    The accelerated growth of algal blooms in water bodies has caused the increased occurrence of taste and odor (T&O) episodes worldwide. Even though T&O compounds have not been associated with adverse health effects, their presence can have extensive socio-economic impacts in contaminated waters. Eagle Creek Reservoir, a eutrophic water body, which supplies about 80% of Indianapolis drinking water, experiences frequent and sometimes severe odorous outbreaks. The terpenoid bacterial metabolites, 2-methylisoborneol (2-MIB) and geosmin, have been identified as the main compounds contributing to those T&O problems, which occur seasonally when the reservoir receives most of its water and nutrient loads from discharge events. In this study, ECR’s microbial community composition was assessed by a 16S next generation sequencing approach, confirming the presence of the major bacterial phyla of Cyanobacteria, Proteobacteria, Actinobacteria and Bacteroidetes, which are commonly found in freshwater environments. The relative abundance of Cyanobacteria, which are regarded as the main T&O producers in freshwater, followed the fluctuation of 2-MIB and geosmin concentrations closely. Mapping sequence analysis of a metagenomic dataset, successfully recovered the genes responsible for the synthesis of geosmin and 2-MIB, demonstrating the microbial ability for odorous compound production in ECR. Quantification of the geoA and MIBS genes in Cyanobacteria was achieved by the development and application of qPCR assays on water samples collected from the reservoir. A statistically significant positive correlation was found between MIBS gene quantity and MIB concentration for all sampling locations, implying that this assay could potentially be used as a tool for the early prediction of upcoming T&O episodes. The geoA gene detection assay, did not correlate well with geosmin concentrations, suggesting that even though the gene might be present, this does not necessarily mean that it is metabolically active.
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    IL-33 Mediated Th2 Effector Functions are Suppressed in Tregs by Bcl6 and Regulated by Sex
    (2024-08) Lee, Kyu Been; Dent, Alexander; Richer, Martin; Robinson, Christopher; Yang, Kai
    Allergic airway inflammation (asthma) is a prevalent and uncurable disease worldwide, affecting many individuals’ quality of life. Although asthma does not form from a singular cause, one primary mediator comes from the exposure to environmental allergens and the improper activation of the T cell subset: T helper 2 (Th2) cells. Th2 cells produce pro-inflammatory cytokines and promote the activation and recruitment of various pro-inflammatory cells into the lung, causing greater damage and inflammatory responses in the organ. Th2 cell’s activation is regulated by another T cell subset, Regulatory T (Treg) cells, by expressing anti-inflammatory cytokines and downregulating the inflammatory response. On the contrary, the release of interleukin-33 (IL-33) from damaged lung epithelial cells transitions Tregs into Th2-like Tregs (ST2+ Tregs) which release both pro-and anti-inflammatory cytokines and cannot suppress the inflammatory disease. However, transcriptional repressor protein B cell lymphoma 6 (Bcl6) provides Tregs a stable follicular phenotype and suppresses the ST2+ Treg transition. Preliminary data revealed that Bcl6 repressive function is dependent on mouse sex, in which Tregs of male mice are more resistant to the ST2+ Treg phenotype than those of female mice. However, the removal of Bcl6 also removed the sex-dependent suppression against the ST2+ Treg transition. The project therefore sought to further confirm and answer whether Bcl6 suppressed the ST2+ Treg phenotype in a sex-dependent manner, ultimately leading to a sex-biased asthma prevalence and severity. We utilized quantitative polymerase chain reaction (qPCR) and next-generation sequencing techniques to uncover which genes Bcl6 regulates, how IL-33 affects chromatin accessibility/gene expression, and what relation sex hormones have with Bcl6 in the expression of Th2 cytokines from Tregs. Currently, we have discovered that estrogen-like chemicals in common cell culturing media may be acting on the estrogen receptor of Tregs and causing differential gene expressions based on media conditions. We also determined that Bcl6 is acting independently of mouse sex to suppress Th2 genes in Tregs, contrary to preliminary findings. Overall, we have obtained insight on the role of the estrogen receptor and Bcl6’s mechanism of suppression in relation to sex.
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    A plasma telomeric cell-free DNA level in unaffected women with BRCA1 or/and BRCA2 mutations: a pilot study
    (Impact Journals, 2017-12-29) Dey, Shatovisha; Marino, Natascia; Bishop, Kanokwan; Dahlgren, Paige N.; Shendre, Aditi; Storniolo, Anna Maria; He, Chunyan; Tanaka, Hiromi; Medical and Molecular Genetics, School of Medicine
    Plasma cell-free DNA (cfDNA) is a small DNA fragment circulating in the bloodstream originating from both non-tumor- and tumor-derived cells. A previous study showed that a plasma telomeric cfDNA level decreases in sporadic breast cancer patients compared to controls. Tumor suppressor gene products including BRCA1 and BRCA2 (BRCA1&2) play an important role in telomere maintenance. In this study, we hypothesized that the plasma telomeric cfDNA level is associated with the mutation status of BRCA1&2 genes. To test this hypothesis, we performed plasma telomeric cfDNA quantitative PCR (qPCR)-based assays to compare 28 women carriers of the BRCA1&2 mutation with age-matched controls of 28 healthy women. The results showed that the plasma telomeric cfDNA level was lower in unaffected BRCA1&2 mutation carriers than in age-matched controls from non-obese women (BMI < 30), while there was no association between unaffected BRCA1&2 mutation carriers and age-matched controls in obese women (BMI > 30). Moreover, the plasma telomeric cfDNA level applied aptly to the Tyrer-Cuzick model in non-obese women. These findings suggest that circulating cfDNA may detect dysfunctional telomeres derived from cells with BRCA1&2 mutations and, therefore, its level is associated with breast cancer susceptibility. This pilot study warrants further investigation to elucidate the implication of plasma telomeric cfDNA levels in relation to cancer and obesity.