S-STEM Scholar Leilani Boren's Blog

 Introduction     Previous tests conducted with qPCR done by myself and Chad Albert have verified that the acylase and lactonase primers that we possess have sufficient efficiency (within 90 to 110 percent) to begin tests evaluating how oxidative stress affects these gene expressions. Among acylase and lactonase there will be four other targets genes tested including Met H, Met K, pfs, Lux S, and two reference genes, secA and GAP3. Deinococcus radiodurans  is significant because it produces both acylase and lactonase, when typically only one is expressed in any given bacteria. By completing this qPCR experiment, a comprehensive understanding of how D. radiodurans responds to oxidative stress will be possible. The preliminary steps to acquire this data will be to inoculate and allow for sufficient growth, then complete a successful RNA extraction from six samples in which half of these samples were exposed to oxidative stress. From these RNA samples, cDNA synthesis wi...

 Introduction     Previous methodology has explained how to take an RNA sample from D. radiodurans and covert that sample into cDNA in order to run qPCR testing to evaluate gene expression. RNA isolation was practiced this week, guided by Kendrick Freeland, with D. radiodurans . Kendrick is currently working on a project that involves using qPCR to observe any differences in gene expression of RecA, ZWF, and GND within D. radiodruans  under oxidative stress compared to no known stressors. The three genes being targeted are all associated with the pentose phosphate pathway, the metabolic pathway that breaks down sugars as an alternative to glycolysis. This pathway also crates free nucleotides as a by product of the process, which is suspected of helping the cells repair damages to their DNA molecules. RecA is known to facilitate the repair of double stranded breaks, ZWF codes for G6PD, an enzyme acting as a limiting factor of the pentose phosphate pathway. GND synthes...

 Introduction     Deinococcus radiodurans  is a gram negative extremophilic bacterial species that is frequently recognized for being able to withstand environmental conditions with high levels of radiation. It is also recently found to be one of the few, potentially only, species that produces both AHL-acylase and AHL-lactonase in order to degrade AHLs. AHL, or N-acyl homoserine lactones, are signal molecules produced by cells, typically in a biofilm community, in order to alter gene expression in the recipient cells. The communication between cells is described as quorum sensing, a survival strategy that regulates the behavior of a community of cells. Most cells that have enzymes that degrade AHLs have either AHL-acylase or AHL-lactonase, but it has been found that Deinococcus radiodurans has genes to synthesize both enzymes, which is incredibly unusual. AHL-acylase is a protein that hydrolyzes the amide bond between the lactone ring and the acyl chain of...