Optimization of Bacterial Bioluminescence (\u3ci\u3elux\u3c/i\u3e) Expression and Development of Autonomous \u3ci\u3elux\u3c/i\u3e-Based Reporters in Human Cell Lines

Despite its extensive use as a prokaryotic bioreporter, only recently has the bacterial bioluminescence (lux) system been adapted to express at a functional level in the mammalian cellular background. While this novel strategy has the potential to contribute to the fields of high throughput screening and non-invasive in vivo imaging due to its autonomous substrate production, it is still constrained because its signal intensity is lower than that of other bioluminescent reporters. This work demonstrates the development of strategies that optimize human cell lux-based bioluminescence to overcome this detriment for advancement towards a fully functional lux reporter system. To enhance lux gene expression, a single vector construct has been developed that improves upon the initial two plasmid expression system. This construct separates the lux genes using viral-derived 2A elements, allowing for simultaneous expression of the six genes from a single promoter in eukaryotic cells. This strategy results in increased bioluminescent output compared to the previous two plasmid system. Additionally, it is demonstrated that gene copy number is the primary limiting factor for bioluminescent output following expression in human cell lines. This limitation has been overcome through the development of a two-step transfection strategy that yields significantly higher transfection efficiency and improves both transgene integration and bioluminescence output. An optimized HEK293 cell line with enhanced bioluminescent production has been constructed using the two-step transfection strategy. The enhanced signal intensity allows bioluminescent detection from a smaller population size in cell culture compared to their un-optimized counterparts, as well as detection using alternative lower cost instruments. A developmental effort towards lux-based biosensing in human cell lines is also reported in this study. It has been shown that bioluminescence emitted constitutively from lux-expressing human cell lines can be utilized for monitoring population dynamics in a non-invasive manner and that, by regulating the expression of the lux genes, the lux system can function as a fully autonomous bioreporter for continuous monitoring of targets of interest

Exploring the Experiences of Male Early Childhood Aspiring Teachers

This exploratory study investigated the reasons why males chose early childhood education and their lived experiences in a teacher education program. Qualitative data were collected through interviews with twelve male early childhood pre-service teachers. Results demonstrated that: Participates were passionate about teaching young children. They were mostly supported by their relatives and friends in their decision to become an early childhood teacher. They were positive about future career opportunities but still had an underlying fear of losing job or being seen as sexual predators. Meanwhile, participates indicated some conflicts might exist between the field experience and program learning experience. They also highlighted a need to recruit male pre-service teachers in early childhood education programs

Using Wireless Pedometers to Measure Children’s Physical Activity: How Reliable is the Fitbit Zip?

The purpose of this study is to examine the reliability of wireless pedometers in measuring elementary school children’s physical activity. Activity measurement using a wireless pedometer Fitbit ZipTM was compared to activity measurement using Yamax Digi-WalkerTM SW701 for a group of randomly selected 25 children in Grades 3, 4, and 5. Fitbit ZipTM wireless pedometers were found to have an appropriate degree (Nunnally & Bernstein, 1994) of accuracy and reliability compared to the Yamax Digi-WalkerTM SW701 pedometer. The Fitbit ZipTM wireless pedometer collected more step counts than the Yamax Digi-WalkerTM SW701 pedometer; however, the difference was not statistically significant. Participants reported that they preferred wearing the Fitbit ZipTM to the Yamax Digi-WalkerTM SW701 because the Fitbit ZipTM was more comfortable to wear and less likely to fall off. Participants also reported being more motivated to move while wearing the Fitbit ZipTM

READY TO LEARN: THE IMPACT OF THE MORNING BLAST PHYSICAL ACTIVITY INTERVENTION ON ELEMENTARY SCHOOL STUDENTS

Objective: The purpose of this study was to investigate the effects of a physical activity intervention programme, named “Morning Blast”, on elementary school students’ math learning and daily physical activity. The Morning Blast intervention programme was a 16-week cardiovascular endurance emphasized physical activity program that students voluntarily participated in before the school day. Participants that volunteered, did so for the duration of the program. Methods: This mixed-methods study included seven educators and 83 students (n=90). The students were all children who were enrolled in Grades 3, 4, and 5 in a semi-rural elementary school in the United States. Data were collected through focus-group interviews, surveys, quantitative analysis of step counts, and from quasi-experimental research design. Results: Students in the experimental group were found to have: (1) increased scores on math standard score, (2) greater confidence in their academic ability, and (3) had more accumulated steps compared to students in the control group. Students in the experimental group also reported that they were more “ready to learn” after completing the physical activity intervention. This finding was also confirmed by their teachers. Conclusion: This study demonstrates how an increase in physical activity during the morning time has positive benefits for students throughout the school day

Applications of Molecular Spectroscopic Methods to the Elucidation of Lignin Structure

Lignin in plant cell wall is a complex amorphous polymer and is biosynthesized mainly from three aromatic alcohols, namely, p-coumaryl, coniferyl, and sinapyl alcohols. This biosynthesis process consists of mainly radical coupling reactions and creates a unique lignin polymer in each plant species. Generally, lignin mainly consists of p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) units and is linked by several types of carbon-carbon (β-β, β-5, β-1, and 5–5) and ether bonds. Due to the structural complexity, various molecular spectroscopic methods have been applied to unravel the aromatic units and different interunit linkages in lignin from different plant species. This chapter is focused on the application of ultraviolet (UV) spectroscopy, Fourier transform infrared (FT-IR) spectroscopy, Fourier transform Raman (FT-Raman) spectroscopy, fluorescence spectroscopy, and nuclear magnetic resonance (NMR) spectroscopy to lignin structural elucidation