DNA Replication

Changes in DNA molecules are common, due to the effects of UV light and other external and internal mutagens. Cells have a variety of repair mechanisms which serve to maintain the accuracy of the genetic code. This activity includes a low-cost, safe and technically feasible experiment, which allows students to observe the effects of UV mutagenesis and DNA photorepair in the halophilc archaeon, Haloferax volcanii. An optional extension links this activity to topics of immediate concern to students – how exposure to UVC light contributes to skin cancer risk and the protective effects of sunscreen. Students design and carry out an experiment to test whether SPF 15 sunscreen increases the lethal exposure time for H. volcanii by a factor of 15. Throughout the activity, discussion questions engage students in actively thinking about the biological phenomena and experimental procedures and analysis. This activity is designed for students in college or university genetics, microbiology, or introductory biology courses as well as in high school honors biology courses. Teachers report that this activity was valuable in helping students understand mutagenesis and photorepair and in developing student skills in designing and analyzing experiments.

Through some unexpected and still-unidentified form of molecular texting, the herpes simplex virus (HSV) induces a hormone-like stimulation of cellular DNA replication in nearby, uninfected cells, according to Peter O'Hare of St. Mary's Medical School, Imperial College in London, United Kingdom, and his collaborators. Details appeared 26 August 2015 in the Journal of Virology (doi:10.1128/JVI.01950–15/).

Science students can benefit from visual aids. In biology lectures, visual aids are usually limited to tables, figures, and PowerPoint presentations. In this IRB-approved study, we examined the effectiveness of the use of five prop demonstrations, three of which are at the intersection of biology and chemistry, in three community college biology courses. We hypothesized that students’ performance on test questions is enhanced by the use of prop demonstrations. Consistent with our hypothesis, we showed that students learn more effectively and perform better on questions that relate to demonstrations than on questions related to lessons that do not have a demonstration component.

Following years of widespread use in business and medical education, the case study teaching method is becoming an increasingly common teaching strategy in science education. However, the current body of research provides limited evidence that the use of published case studies effectively promotes the fulfillment of specific learning objectives integral to many biology courses. This study tested the hypothesis that case studies are more effective than classroom discussions and textbook reading at promoting learning of key biological concepts, development of written and oral communication skills, and comprehension of the relevance of biological concepts to everyday life. This study also tested the hypothesis that case studies produced by the instructor of a course are more effective at promoting learning than those produced by unaffiliated instructors. Additionally, performance on quantitative learning assessments and student perceptions of learning gains were analyzed to determine whether reported perceptions of learning gains accurately reflect academic performance. The results reported here suggest that case studies, regardless of the source, are significantly more effective than other methods of content delivery at increasing performance on examination questions related to chemical bonds, osmosis and diffusion, mitosis and meiosis, and DNA structure and replication. This finding was positively correlated to increased student perceptions of learning gains associated with oral and written communication skills and the ability to recognize connections between biological concepts and other aspects of life. Based on these findings, case studies should be considered as a preferred method for teaching about a variety of concepts in science courses.

Stem cells hold great promise in the treatment of diseases ranging from cancer to dementia. However, as rapidly as the field of stem cell biology has emerged, heated political debate has followed, scrutinizing the ethical implications of stem cell use. It is therefore imperative to promote scientific literacy by educating students about stem cell biology. Yet, there is a definite lack of material to engage students in this subject at the basic science level. Therefore, we have developed and implemented a hands-on introductory laboratory module that introduces students to stem cell biology and can be easily incorporated into existing curricula. Students learn about stem cell biology using an in vivo planarian model system in which they down-regulate two genes important in stem cell differentiation using RNA interference and then observe the regenerative phenotype. The module was piloted at the high school, community college, and university levels. Here, we report that introductory biology students enrolled at a community college were able to demonstrate gains in learning after completion of a one-hour lecture and four 45-minute laboratory sessions over the course of three weeks. These gains in learning outcomes were objectively evaluated both before and after its execution using a student quiz and experimental results. Furthermore, students’ self-assessments revealed increases in perceived knowledge as well as a general interest in stem cells. Therefore, these data suggest that this module is a simple, useful way to engage and to teach students about stem cell biology.

Homologous recombination is an ubiquitous process that shapes genomes and repairs DNA damage. The reaction is classically divided into three phases: presynaptic, synaptic, and postsynaptic. In Escherichia coli, the presynaptic phase involves either RecBCD or RecFOR proteins, which act on DNA double-stranded ends and DNA single-stranded gaps, respectively; the central synaptic steps are catalyzed by the ubiquitous DNA-binding protein RecA; and the postsynaptic phase involves either RuvABC or RecG proteins, which catalyze branch-migration and, in the case of RuvABC, the cleavage of Holliday junctions. Here, we review the biochemical properties of these molecular machines and analyze how, in light of these properties, the phenotypes of null mutants allow us to define their biological function(s). The consequences of point mutations on the biochemical properties of recombination enzymes and on cell phenotypes help refine the molecular mechanisms of action and the biological roles of recombination proteins. Given the high level of conservation of key proteins like RecA and the conservation of the principles of action of all recombination proteins, the deep knowledge acquired during decades of studies of homologous recombination in bacteria is the foundation of our present understanding of the processes that govern genome stability and evolution in all living organisms.

This review illustrates the logic style that Roth taught in investigation of chromosome rearrangements and highlights the interesting findings. A few explanations were proposed for the rarity of inversions: inverse-order repeats occur infrequently in the chromosomes of these organisms; inversion rearrangements may cause deleterious effects that cause a selective disadvantage; and mechanistic constraints may prevent the formation of the rearrangements. Inversions were frequently recovered among recombinants when the repeated sequences flanked certain chromosome segments, termed “permissive". The author focuses on the Roth lab’s approach to studying the barriers to inversion of nonpermissive chromosome segments in S. Typhimurium and will correlate findings with outcomes of investigations in E. coli. Alternatively, barriers to the recombination events required for inversion may block formation of the rearrangements. The crosses yielded inversion rearrangements at frequencies expected of two-fragment transduction events. The role of DNA replication is considered in the homologous recombination events that form inversion rearrangements. Investigation of inversion formation by homologous recombination in E. coli found that replication pausing at inverted ter sites is not the only explanation for nonpermissive and restrictive intervals. It remains to be determined if other barriers limit inversion in Salmonella. The findings summarized here highlight topics to investigate regarding the mechanism of inversion formation and barriers to this process for nonpermissive chromosomal segments. The role of DNA replication in inversion formation could be examined by testing the outcome of placing blocked ter sites within permissive arcs of the chromosome.

DNA and RNA helicases are organized into six superfamilies of enzymes on the basis of sequence alignments, biochemical data, and available crystal structures. DNA helicases, members of which are found in each of the superfamilies, are an essential group of motor proteins that unwind DNA duplexes into their component single strands in a process that is coupled to the hydrolysis of nucleoside 5'-triphosphates. The purpose of this DNA unwinding is to provide nascent, single-stranded DNA (ssDNA) for the processes of DNA repair, replication, and recombination. Not surprisingly, DNA helicases share common biochemical properties that include the binding of single- and double-stranded DNA, nucleoside 5'-triphosphate binding and hydrolysis, and nucleoside 5'-triphosphate hydrolysis-coupled, polar unwinding of duplex DNA. These enzymes participate in every aspect of DNA metabolism due to the requirement for transient separation of small regions of the duplex genome into its component strands so that replication, recombination, and repair can occur. In Escherichia coli, there are currently twelve DNA helicases that perform a variety of tasks ranging from simple strand separation at the replication fork to more sophisticated processes in DNA repair and genetic recombination. In this chapter, the superfamily classification, role(s) in DNA metabolism, effects of mutations, biochemical analysis, oligomeric nature, and interacting partner proteins of each of the twelve DNA helicases are discussed.

This review describes the components of the Escherichia coli replisome and the dynamic process in which they function and interact under normal conditions. It also briefly describes the behavior of the replisome during situations in which normal replication fork movement is disturbed, such as when the replication fork collides with sites of DNA damage. E. coli DNA Pol III was isolated first from a polA mutant E. coli strain that lacked the relatively abundant DNA Pol I activity. Further biochemical studies, and the use of double mutant strains, revealed Pol III to be the replicative DNA polymerase essential to cell viability. In a replisome, DnaG primase must interact with DnaB for activity, and this constraint ensures that new RNA primers localize to the replication fork. The leading strand polymerase continually synthesizes DNA in the direction of the replication fork, whereas the lagging-strand polymerase synthesizes short, discontinuous Okazaki fragments in the opposite direction. Discontinuous lagging-strand synthesis requires that the polymerase rapidly dissociate from each new completed Okazaki fragment in order to begin the extension of a new RNA primer. Lesion bypass can be thought of as a two-step reaction that starts with the incorporation of a nucleotide opposite the lesion, followed by the extension of the resulting distorted primer terminus. A remarkable property of E. coli, and many other eubacterial organisms, is the speed at which it propagates. Rapid cell division requires the presence of an extremely efficient replication machinery for the rapid and faithful duplication of the genome.

DNA replication is a topic usually presented in 9th-grade biology. The essential fact of DNA replication is that the base-pairing rules make it very easy to generate two identical new helices from one helix. The first part is appropriate for young students; more advanced students will perform both parts of the lesson. The first activity described in this chapter is a simple (and necessarily inaccurate) paper simulation of DNA replication. The second activity is a student reading about DNA polymerase, the central DNA replication enzyme. The background information in the introduction that follows contains far more detail about DNA replication. The two aspects of DNA synthesis that your advanced students need to know are that synthesis is unidirectional and that it absolutely requires a primer. Not surprisingly, the characteristics of DNA polymerase determine the overall features of DNA replication inside the cell and in the test tube. The feature of DNA replication means that DNA synthesis is unidirectional, from 5' to 3'. Uni-directionality presents a problem for chromosome replication that is discussed in this chapter. Chromosomal DNA replication is usually initiated at specific sites along the DNA called replication origins. Instead, several different strategies circumvent the problem created by the specificity of DNA replication enzymes.