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Research in microbial biofuels has dramatically increased over the last decade. The bulk of this research has focused on increasing the production yields of cyanobacteria and algal cells and improving extraction processes. However, there has been little to no research on the potential impact of viruses on the yields of these phototrophic microbes for biofuel production. Viruses have the potential to significantly reduce microbial populations and limit their growth rates. It is therefore important to understand how viruses affect phototrophic microbes and the prevalence of these viruses in the environment. For this study, phototrophic microbes were grown in glass bioreactors, under continuous light and aeration. Detection and quantification of viruses of both environmental and laboratory microbial strains were measured through the use of a plaque assay. Plates were incubated at 25º C under continuous direct florescent light. Several environmental samples were taken from Tempe Town Lake (Tempe, AZ) and all the samples tested positive for viruses. Virus free phototrophic microbes were obtained from plaque assay plates by using a sterile loop to scoop up a virus free portion of the microbial lawn and transferred into a new bioreactor. Isolated cells were confirmed virus free through subsequent plaque assays. Viruses were detected from the bench scale bioreactors of Cyanobacteria Synechocystis PCC 6803 and the environmental samples. Viruses were consistently present through subsequent passage in fresh cultures; demonstrating viral contamination can be a chronic problem. In addition TEM was performed to examine presence or viral attachment to cyanobacterial cells and to characterize viral particles morphology. Electron micrographs obtained confirmed viral attachment and that the viruses detected were all of a similar size and shape. Particle sizes were measured to be approximately 50-60 nm. Cell reduction was observed as a decrease in optical density, with a transition from a dark green to a yellow green color for the cultures. Phototrophic microbial viruses were demonstrated to persist in the natural environment and to cause a reduction in algal populations in the bioreactors. Therefore it is likely that viruses could have a significant impact on microbial biofuel production by limiting the yields of production ponds.
ContributorsKraft, Kyle (Author) / Abbaszadegan, Morteza (Thesis advisor) / Alum, Absar (Committee member) / Fox, Peter (Committee member) / Arizona State University (Publisher)
Created2014
Description
To identify genes that can lead to obesity of Pima Native American heritage, an array of experiments can be conducted to determine possible candidate genes that can increase the likelihood of being obese in a set population. The studies available to identify these genes were (1) inspect follow-up genes identified by a previous genome wide associations studies, GWAS, previously conducted for the 1120 American Indian subjects data available, (2) to directly sequence candidate genes in literature, (3) to analyze whole sequence data from Native American subjects, and lastly (4) to perform functional studies on most promising variants associated with BMI. Analyzing the results presented from my work required the use of biological techniques such as: DNA sequencing, DNA large scale genotyping, PCR amplification, DNA transfections, DNA ligations, in vitro Luciferase assay and Cell culture. Inspecting the follow-up genes identified by the conducted GWAS showed the potential for the MAP2K3 gene to be a candidate to increase obesity in the set population, involve two single nucleotide polymorphisms (SNPs, rs12882548, rs11652094), to affect body weight through complex mechanisms involving food intake and hypothalamic inflammation. The follow-up genes identified in the GWAS that had an effect on obesity showed to affect it through the mechanism of reducing energy expenditure. Through the analysis of SNPs two variants (rs10507100 and rs17087518) were identified to test their roles in the reduction of energy expenditure. Rs17087518 showed to have a role in a relatively reduced EE resulting in weight gain. Directly sequencing a candidate gene known as MRAP2 showed that the SNP rs1928281 did not have a significant difference on obesity in the Native American subjects (p =.09). Analyzing whole genome sequencing SNPs gave rise to novel variants by association analyses with energy expenditure and BMI in 235 whole genomes, the most significant SNP, rs4984683, was examined to determine the variability in energy expenditures. With set quality control assessment a list of variants were received and were then later assessed with other data available to make a connection to EE. Performing functional studies showed the possibility for rs2001651 and rs1466314 to have an effect on MAP2K3 expression level. The initial functional studies gave way to a more in-depth study of this gene to predict BMI in Caucasians and Native Americans, which in turn showed an association with BMI. The use of these techniques have been an indicator for current research in the determination of candidate genes across many diseases. The works presented is an example of the current works in genetics and an exploration of new mechanism to detect, and possibly treat, disease through personalized sequencing.
ContributorsGale, Alex Mauricio Pompa (Author) / Ankeny, Casey (Thesis director) / Baier, Leslie (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2015-05
Description
This project aims to address the current protocol regarding the diagnosis and treatment of traumatic brain injury (TBI) in medical industries around the world. Although there are various methods used to qualitatively determine if TBI has occurred to a patient, this study attempts to aid in the creation of a system for quantitative measurement of TBI and its relative magnitude. Through a method of artificial evolution/selection called phage display, an antibody that binds highly specifically to a post-TBI upregulated brain chondroitin sulfate proteoglycan called neurocan has been identified. As TG1 Escheria Coli bacteria were infected with KM13 helper phage and M13 filamentous phage in conjunction, monovalent display of antibody fragments (ScFv) was performed. The ScFv bind directly to the neurocan and from screening, phage that produced ScFv's with higher affinity and specificity to neurocan were separated and purified. Future research aims to improve the ScFv characteristics through increased screening toward neurocan. The identification of a highly specific antibody could lead to improved targeting of neurocan post-TBI in-vivo, aiding researchers in quantitatively defining TBI by visualizing its magnitude.
ContributorsSeelig, Timothy Scott (Author) / Stabenfeldt, Sarah (Thesis director) / Ankeny, Casey (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2015-05