Establishing live-cell single-molecule localization microscopy imaging and single-particle tracking in the archaeon Haloferax volcanii

In recent years, fluorescence microscopy techniques for the localization and tracking of single molecules in living cells have become well-established and indispensable tools for the investigation of cellular biology and in vivo biochemistry of many bacterial and eukaryotic organisms. Nevertheless, these techniques are still not established for imaging archaea. Their establishment as a standard tool for the study of archaea will be a decisive milestone for the exploration of this branch of life and its unique biology. Here we have developed a reliable protocol for the study of the archaeon Haloferax volcanii. We have generated an autofluorescence-free H. volcanii strain, evaluated several fluorescent proteins for their suitability to serve as single-molecule fluorescence markers and codon-optimized them to work under optimal H. volcanii cultivation conditions. We found that two of them, Dendra2Hfx and PAmCherry1Hfx, provide state-of-the-art single-molecule imaging. Our strategy is quantitative and allows dual-color imaging of two targets in the same field of view as well as DNA co-staining. We present the first single-molecule localization microscopy (SMLM) images of the subcellular organization and dynamics of two crucial intracellular proteins in living H. volcanii cells, FtsZ1, which shows complex structures in the cell division ring, and RNA polymerase, which localizes around the periphery of the cellular DNA. This work should provide incentive to develop SMLM strategies for other archaeal organisms in the near future.</jats:p

Investigation of eighth-grade students' understanding of the slope of the linear function

This study aimed to investigate eighth-grade students' difficulties and misconceptions and their performance of translation between the different representation modes related to the slope of linear functions. The participants were 115 Turkish eighth-grade students in a city in the eastern part of the Black Sea region of Turkey. Data was collected with an instrument consisting of seven written questions and a semi-structured interview protocol conducted with six students. Students' responses to questions were categorized and scored. Quantitative data was analyzed using the SPSS 17.0 statistical packet program with cross tables and one-way ANOVA. Qualitative data obtained from interviews was analyzed using descriptive analytical techniques. It was found that students' performance in articulating the slope of the linear function using its algebraic representation form was higher than their performance in using transformation between graphical and algebraic representation forms. It was also determined that some of them had difficulties and misunderstood linear function equations, graphs, and slopes and could not comprehend the connection between slope and the x- and y-intercepts

Structural Basis for the Aminoacid Composition of Proteins from Halophilic Archea

In order to survive in highly saline environments, proteins from halophilic archea have evolved with biased amino acid compositions that have the capacity to reduce contacts with the solvent