The activity and expression levels of tripeptidyl-peptidase II (TPP-II), a proteolytic protein complex, is unknown in diabetic muscle tissue. To investigate TPP-Iwestern blotting and biological assays were employed to answer the questions: 1) Do TPP-II levels differ in Akita mice versus wild-type mice? and 2) Is TPP-II activity different in Akita and wild-type mice? Before responding to these questions, however, the common western blotting technique used was optimized to improve the sensitivity of the blot. The optimal conditions for western blotting were determined to be 5% PEG-8000 + 0.01% Glutaraldehyde + 75% TTBS and overnight primary antibody incubation. Using this optimized western blotting, TPP-II levels were determined to be the same in Akita and wild-type mice. TPP-II activity was also determined and found to be similar between hearts from Akita and wild-type mice.
The ability to form complex memories is a well studied yet not well understood mechanism of the brain. Several studies have been performed on the creation of episodic memories from object representations, but none have examined the effect of multiple types of contextual cues and how they are prioritized during memory encoding. In this study, we examined the creation of episodic memories via the integration of multiple different elements. Sixty-four subjects, all of normal health, navigated a system of virtual reality rooms and answered questions about the room and music’s emotional effect during a study phase. During the testing phase, they once again navigated the hallway but answered questions that tested for recall of details in rooms that they entered. We then quantified this data by counting corrects versus false alarms and analyzed the data using a one-way ANOVA. We examined three model effects: delay between the study and test phases, the subjects’ type of recall – spatial, musical, or both – as determined by the questions, and the interaction between delay and recall type. Our findings indicate that music, as a lone contextual cue, provides an advantage over spatial cues during encoding that later presented itself in retrieval. However, this effect dissipated after one week. It therefore appears that the impact of varying types of contextual details changes differently over time, although a combination of multiple details is always better than a single type of detail.
The effect of microwave intensity and exposure on the carotenoid concentration of a cassava gari meal with added red palm oil.
Cassava gari is a staple food in Africa, which has been biofortified to help combat vitamin A deficiency. However, a correlation is often seen between the extent of gari preparation and a decrease in the carotenoid concentration of the gari. The objective is to find out if higher microwave intensities and longer duration of exposure to microwaves may decrease the carotenoid concentration in cassava gari.
Spatio-temporal dynamics and physiological characterization of post harvest chilling in tomato fruit.
Magnetic resonance imaging was used to spatially and temporally examine physiological changes in tomato fruit (S. lycopersicum var. cerasiforme cv. sweet 100) caused by exposure to chilling temperatures. Measurements for respiration and ethylene production showed the development of chilling injury as indicated by the chilling injury index. The MRI was used to calculate the apparent diffusion coefficient (ADC), a measure of water mobility in tissues, for different regions of interest (ROI) in the fruit.
Small unilamellar vesicles (SUVs) aptly model cell membranes. In fact, vesicles are currently a major area of focus. They are being used to study biological functions through the incorporation of various molecules into the vesicle membrane . In this research, the structure of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) 30 and 100 nanometer vesicles were studied in phosphate buffered saline (PBS) and water. The lipid samples were adsorbed on an oxidized gold surface on a germanium crystal and studied with attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR). In both PBS and water, 30 and 100 nm vesicles adsorbed intact. However, deformation of the SUVs varied with the size of the vesicles and solvent. This study shows that DMPC vesicles reacted differently based on changes to these two factors. These observations will help advance surface chemistry research and the development of better cell bilayer models.
The posttranslational modification of tyrosine sulfation is critical for the binding affinity and peptide specificity of many G-protein coupled receptors (GPCRs). Using a log-odds position specific scoring matrix (PSSM), 64 tyrosine sulfation sites in 31 different peptide binding GPCRs were scored. Higher scores indicated a higher degree of similarity between confirmed sulfation sites and that tyrosine. While our PSSM did not account for characteristics such as clustering and conservation, the sites nonetheless exhibited these characteristics. Most predicted sites were located in the binding pockets of GPCRs, which is also consistent with confirmed tyrosine sulfation sites. Binding affinity or peptide specificity decreased when our tyrosines were mutated into alanine or phenylalanine, indicating the important role of the tyrosine in ligand binding.
In N.tabacum, chloroplastic protein NRIP1 mediates the innate immune receptor recognition of p50, the viral effector protein of the Tobacco Mosaic Virus. The presence of stromules was observed extending from the stroma of the chloroplasts towards the nucleus. Researchers discovered that ferritin, another chloroplastic protein, does not complete a similar process to NRIP1, instead, it is always located in small quantities in the nucleus that do not appear to change during N-mediated defense.
The goal of this project was to determine if Glutathione S-Transferase Mu 1 (GSTM1), a key antioxidant enzyme, is altered in diabetes. We compared GSTM1 levels between hearts of type I diabetic Akita mice and wild-type mice using Western blotting. Western blotting for GSTM1 had previously been unsuccessful because of the low amounts of this enzyme in the heart. The Western blotting procedure was optimized through experiments to determine if treatment of the membrane prior to blocking and during primary and secondary antibody incubation increased the sensitivity of the blot. We found that treatment of the membrane in 0.01% glutaraldehyde for 20 minutes before blocking and incubating with the primary antibody in the presence of 75% TTBS (Tris-buffered saline with Tween 20) increased band intensity by 8.92-fold when compared to the control. In addition, the addition of 5% PEG-8000 and the use of 1% BSA in the primary and secondary antibody incubations increased Western blot sensitivity. The improved Western blotting procedure allowed for the detection and quantification of the GSTM1 in the Akita and control samples. After quantifying and normalizing the protein loaded, we found that GSTM1 levels were similar in Akita mice than in normal mice. Hence, the levels of GSTM1 do not seem to be affected in type 1 diabetic heart dysfunctions.
Greenhouse studies were performed to examine the effects of two carbon sources on Anaerobic Soil Disinfestation (ASD) of Agrobacterium tumefaciens and Pseudomonas synxantha inoculum. The first experiment was performed using Yolo silty clay loam soil from the UC Davis Armstrong Field Facility, which will be referred to as clay soil. The experimental design was composed of treatment pots in a controlled greenhouse environment, each with an irrigated soil amended with rice bran at a rate of 9, 7, or 5 tons/acre, or molasses at a rate of 4.5, 2.25, or 1.125 tons/acre. Inoculum bags containing the bacteria A. tumefaciens and P. synxantha were buried in these pots, which were then covered with an impermeable tarp. The second experiment only considered the treatment of 9 tons/acre of rice bran, but used soil with a Hanford fine sandy loam texture, which will be referred to as sandy soil. Untreated control pots were also present for comparison in both experiments. At three and seven days after initiating Anaerobic Soil Disinfestation, three pots from each treatment were taken to enumerate the bacteria through dilution plating on selective media. Populations of A. tumefaciens in the 1.125 tons/acre molasses treatment tended to decrease with time, but the populations at seven days were not significant different from time zero across treatments. Populations of P. synxantha in the 9 tons/acre rice bran treatment also tended to decrease with time, but the populations were also not significantly different from time zero across treatments. Anaerobic conditions were lost after five days for both the 1.125 tons/acre molasses 2.25 tons/acre molasses, with lower efficacy of ASD over time for molasses compared to the rice bran treatments. A. tumefaciens populations decreased to undetectable levels by seven days in the sandy soils exposed to ASD conditions. These data indicate that soil texture may play a major role in ASD efficacy which will affect where ASD will be used commercially in the state of California. Overall, the comparison of rice bran and molasses as carbon sources for ASD was inconclusive; however, it was clearly shown that rice bran at rates below commercial levels of 9 tons/acre were effective at inducing anaerobic conditions in the soil.
Refining the metagenomic approach used to study sanitizer-treated sludge microbiota and orange peels with limonene
To maximize biofuel methane production from food processor waste, our research sought to develop a system that could be used to genetically examine methanogens that are resistant to industrial and natural sanitizers. We especially focused on the orange juice processor waste, which contained the natural sanitizer limonene and the protein pectin that inhibited DNA extraction. Hermetic bioreactors filled with tomato pomace and sludge were connected to a respirometer, which quantified the gases they released. After testing factors such as the nutrient medium, it was found that higher volumes (>100 mL) of brand new sludge taken straight from refrigeration was needed for reliable results. The results of the following test trials showed that methanogens were increasingly hindered by increasing sanitizer concentrations above 30 mg/L, signifying that the system functioned properly. In another experiment, DNA was best extracted from a mixture of orange peels and limonene by lysing the samples without centrifuging or bead beating and using pectinase to digest the pectin. The concentration of DNA was qualitatively analyzed using spectrophotometry and Qubit®. For polymerase chain reaction (PCR) amplification, bovine serum albumin (BSA) was used to enhance the compatibility of the DNA strand as well as to prevent inhibitors from interfering with the process. Such a systematic procedure could not only be utilized to discover the genes coding for sanitizer resistance, but also potentially be applied to test a wide variety of variables that may affect methane generation.