Utilization of Escherichia Coli for the growth of Y Family DNA Polymerase Rev1 and GSTrap column for purification​

Rev1 is a Y family DNA polymerase that specializes in translesion DNA synthesis. Rev1 is unique in that it preferentially incorporates dCTP in the growing DNA strand, regardless of the templating base. This is because the template base is evicted from the active site and a template amino acid, arginine 324 (R324) acts as the template for the incoming dCTP. We hypothesize that arginine 324 and the neighboring leucine (L325) facilitate the eviction of the DNA template from the active site. To test this hypothesis, we worked to purify R324G/L325G Rev1 double mutant for the purpose of X-ray crystallographic examination of the protein-DNA-dNTP ternary complex. We transformed Escherichia coli (E. coli) and induced expression of both wild type Rev1 and R324G/L325G Rev1. The bacterial cells were lysed by sonification, and the lysate was purified with a GSTrap column. We were able to successfully isolate the Rev1 enzyme. Further purification and crystallization will be necessary to explore the x-ray crystal structure of R324G/L325G Rev1 protein

Genetic Annotation of Bacteriophages MScarn, Knocker, and Neos5

We annotated the genomes of three recently discovered bacteriophages to learn more about their genetic composition. MScarn is a lytic bacteriophage that infects Gordonia terrae 3612. It was discovered and purified from soil collected in Iroquois, SD. MScarn is a cluster CT phage, one of only 37 discovered to date. Its genome is 45,677 base pairs long and has 10-nucleotide 3’ sticky overhanging ends. Its GC content is 60.3% which is typical of CT cluster members. Knocker is a cluster B9 phage that was isolated on the host Mycobacterium smegmatis mc²155 from soil collected in Watertown, SD. Its circularly permuted genome contains 71,459 base pairs, and it has a high GC content of 69.7%. Similar to the other three members of the B9 cluster, it exhibits a lytic life cycle. Neos5, a lytic bacteriophage, was also isolated on Mycobacterium smegmatis mc²155 from soil collected in Baltimore, MD. It is a cluster B3 phage with a circularly permuted genome of 68,886 base-pairs and a 67.5% GC content, synonymous to the other 37 members of the cluster. All three phages were discovered, purified, and annotated by Northwestern College students

Understanding the Antiproliferative Activity of Plant Extracts

Many plants possess medicinal properties. Some, such as the Pacific yew, have yielded chemotherapeutic drugs (taxanes). Scientists report that other extracts such as the leaves of Calendula officinalis (marigold), Vinca rosea (periwinkle), Viscum cruciatum (mistletoe), and Rosmarinus officinalis (rosemary) have anti-tumor activity. In most cases, the chemical components responsible for antiproliferative activity have not been identified and it is unclear if any individual components are as effective in isolation as they are in the context of the whole extract. Furthermore, in most cases, there are no data indicating whether these extracts have synergistic effects or cause negative reactions when used with other drugs. We are using HeLa (adenocarcinoma), RAW 264.7 (leukemia), HepG2 (hepatoma), MDA-MB-231 (adenocarcinoma), and human foreskin fibroblasts (HFF, non-tumorigenic) to test the antiproliferative activity of several plant extracts. We identified five extracts, grapeseed, guava, yew, juniper berry, and Vinca, that slow the growth of all five cell lines in a dose-dependent manner. We are using a variety of methods to understand the mechanism by which these extracts are blocking cell growth

Black Box 2020

The Black Box is a student based creative publication serving the Embry-Riddle Prescott campus. It is our goal to provide a showcase for the creativity and talent of the Embry-Riddle students. Creative works by members of Embry-Riddle\u27s faculty and staff are included

Genomic Annotation of Bacteriophages Clayda5, GShelby23, Santhid, and Wrigley

We annotated the genomes of four recently discovered Actinobacteriophages. Clayda5 and GShelby23 were isolated on Microbacterium foliorum NRRL B-24224. Clayda5 is a lytic, cluster EB phage, one of only 47 discovered to date. It has 10 base pair 3’ sticky overhanging ends and a GC content is 67.2%. It has 70 protein-coding genes and two tRNA genes in its 39,894 bp genome. Clayda5 was purified from soil collected in Hull, IA. GShelby23 was isolated from soil collected in Storm Lake, IA. It is a cluster EM phage, one of only six discovered to date. Its genome is circularly permuted and 53,603 bp long. Its GC content is 64.8%. Santhid and Wrigley are phages that infect Gordonia terrae 3612. Santhid is a cluster DY phage, one of only five discovered to date. It was isolated from soil collected in Orange City, IA. Its genome is 39,295 bp long and includes 60 protein-coding genes. Its GC content is 67.7% and has 10 base pair 3’ sticky overhanging ends. Wrigley was isolated using an enrichment protocol from soil collected in Johnston, IA. It is a cluster CY phage, one of only 17 discovered to date. It is a temperate phage whose genome is 51,878 bp long and includes 81 protein-coding genes. It has 10 base pair 3’ sticky overhanging ends and a GC content of 66.3%.

A systems approach to assess climate change mitigation options in landscapes of the United States forest sector

Abstract Background United States forests can contribute to national strategies for greenhouse gas reductions. The objective of this work was to evaluate forest sector climate change mitigation scenarios from 2018 to 2050 by applying a systems-based approach that accounts for net emissions across four interdependent components: (1) forest ecosystem, (2) land-use change, (3) harvested wood products, and (4) substitution benefits from using wood products and bioenergy. We assessed a range of land management and harvested wood product scenarios for two case studies in the U.S: coastal South Carolina and Northern Wisconsin. We integrated forest inventory and remotely-sensed disturbance data within a modelling framework consisting of a growth-and-yield driven ecosystem carbon model; a harvested wood products model that estimates emissions from commodity production, use and post-consumer treatment; and displacement factors to estimate avoided fossil fuel emissions. We estimated biophysical mitigation potential by comparing net emissions from land management and harvested wood products scenarios with a baseline (‘business as usual’) scenario. Results Baseline scenario results showed that the strength of the ecosystem carbon sink has been decreasing in the two sites due to age-related productivity declines and deforestation. Mitigation activities have the potential to lessen or delay the further reduction in the carbon sink. Results of the mitigation analysis indicated that scenarios reducing net forest area loss were most effective in South Carolina, while extending harvest rotations and increasing longer-lived wood products were most effective in Wisconsin. Scenarios aimed at increasing bioenergy use either increased or reduced net emissions within the 32-year analysis timeframe. Conclusions It is critical to apply a systems approach to comprehensively assess net emissions from forest sector climate change mitigation scenarios. Although some scenarios produced a benefit by displacing emissions from fossil fuel energy or by substituting wood products for other materials, these benefits can be outweighed by increased carbon emissions in the forest or product systems. Maintaining forests as forests, extending rotations, and shifting commodities to longer-lived products had the strongest mitigation benefits over several decades. Carbon cycle impacts of bioenergy depend on timeframe, feedstocks, and alternative uses of biomass, and cannot be assumed carbon neutral

Coronal Heating as Determined by the Solar Flare Frequency Distribution Obtained by Aggregating Case Studies

Flare frequency distributions represent a key approach to addressing one of the largest problems in solar and stellar physics: determining the mechanism that counter-intuitively heats coronae to temperatures that are orders of magnitude hotter than the corresponding photospheres. It is widely accepted that the magnetic field is responsible for the heating, but there are two competing mechanisms that could explain it: nanoflares or Alfv\'en waves. To date, neither can be directly observed. Nanoflares are, by definition, extremely small, but their aggregate energy release could represent a substantial heating mechanism, presuming they are sufficiently abundant. One way to test this presumption is via the flare frequency distribution, which describes how often flares of various energies occur. If the slope of the power law fitting the flare frequency distribution is above a critical threshold, $\alpha=2$ as established in prior literature, then there should be a sufficient abundance of nanoflares to explain coronal heating. We performed $>$600 case studies of solar flares, made possible by an unprecedented number of data analysts via three semesters of an undergraduate physics laboratory course. This allowed us to include two crucial, but nontrivial, analysis methods: pre-flare baseline subtraction and computation of the flare energy, which requires determining flare start and stop times. We aggregated the results of these analyses into a statistical study to determine that $\alpha = 1.63 \pm 0.03$. This is below the critical threshold, suggesting that Alfv\'en waves are an important driver of coronal heating.Comment: 1,002 authors, 14 pages, 4 figures, 3 tables, published by The Astrophysical Journal on 2023-05-09, volume 948, page 7