bluuebunny/crossref_metadata_2025 huggingface dataset:
Development of a biochemistry laboratory course with a project‐oriented goal
Authors: Stahelin Robert V. , Forslund Raymond E. , Wink Donald J. , Cho Wonhwa
Published: February 2007

A new course for undergraduate biochemistry laboratory has been developed in the Department of Chemistry at the University of Illinois at Chicago. The course is innovative pedagogically by giving students a set of relevant skills for biochemical research and then allowing them to apply those skills in designing and performing the procedures for a research‐like project. The course content focuses on the pharmacologically important enzyme β‐lactamase, which is involved in the hydrolysis of antibiotics such as penicillins and cephalosporins. Specifically, each student designs and performs site‐directed mutagenesis of the enzyme β‐lactamase and analyzes the effect of mutation on the catalytic function of the enzyme by an enzyme assay. This unique and valuable research experience greatly enhances their understanding of scientific reasoning and the research process. Evaluation of the progress also indicates successful linkage of skill‐building and student‐directed activities even for students with no prior experience.
Different energy sources in sports: Introductory software
Authors: Galembeck Eduardo , Macedo Denise V. , Torres Bayardo B.
Published: February 2007

This work describes software designed to promote the association between the content of a basic undergraduate biochemistry course and the professional activities of physical education students. The software contains three main content sections: (A) Structure of skeletal muscle, (B) Contraction mechanism, and (C) Adaptations to physical exercise. A fourth section asks questions of the students. The software offers students a brief introduction on muscle structure and function, focusing on the energy sources required by different kinds of physical activities. The software was field tested at Universidade Estadual de Campinas (Unicamp), Brazil and has been adopted by several Brazilian universities. The students are required to examine the software, to discuss its contents, and to produce a list of questions arising from their work with the software. These questions are answered during the development of the curriculum, thereby connecting biochemical knowledge with the energetic needs for the practice of sports.
Content and conflict: The use of current events to teach content in a biochemistry course*
Authors: Cornely Kathleen
Published: February 2007

Case study exercises, based on newsworthy current events, have been written for a biochemical audience that include elements of both “content” and “conflict.” The cases have been written in such a way as to teach students basic biochemical principles. At the same time, a dilemma is posed to the students, who must use what they have learned about the issue to come to a decision to resolve the conflict. In this manner, the students learn important biochemical principles but are also asked to deal with the scientific or social conflict posed in the case.
Energy diagrams for enzyme‐catalyzed reactions: Concepts and misconcepts
Authors: Aledo J. Carlos , Lobo Carolina , del Valle Alicia Esteban
Published: February 2007

Despite the utility that energy diagrams have as a teaching and learning tool, a survey of their use, in seven popular Biochemistry textbooks, reveals that there is certain confusion around this topic. In our opinion, this confusion arises from the reluctance of authors to consider and indicate the conditions under which the reaction being represented occurs. For an enzyme‐catalyzed reaction, it should be stressed that, under conditions where the overall reaction is spontaneous, each elementary step must exhibit a negative free energy change, and this must be properly reflected in the progression profile of the reaction.
Isolation, purification, and characterization of bovine heart and yeast cytochromes c: An integrated biochemistry laboratory experience
Authors: Hunnes Cristi H.
Published: February 2007

This article describes a one‐semester project‐based laboratory course for biochemistry students centered on the protein cytochrome c. Students learn a variety of protein purification techniques, including ammonium sulfate precipitation, dialysis, ultrafiltration, gel filtration chromatography, ion‐exchange chromatography, and electrophoresis, by isolating cytochrome c. The oxidation‐reduction properties of this relatively stable and forgiving protein are then characterized by UV‐visible spectroscopy and electrochemistry. Electrophoresis is utilized to assess purity and determine molecular weight. Students isolate cytochrome c from bovine heart and yeast and compare their properties.
Disciplinary differences or modes of curriculum practice? Who promised to deliver what in problem‐based learning?
Authors: Savin‐Baden Maggi
Published: February 2007

Although problem‐based learning has become increasingly popular in the last five years or so, particularly in the United Kingdom, the arguments about it still seem to rage. The ones that seem to be most problematic are, first, those about the relationship between problem‐based learning and other similar active learning approaches, such as project‐based learning; and second, those arguments about sustaining problem‐based learning curricula in the face of cuts and poor university administration. Meanwhile, the blame‐it‐on‐problem‐based‐learning attitude still seems to prevail worldwide, with little real consideration of the broader concerns about some of the educational issues. For example, there seem to be too few debates about what it means to facilitate problem‐based learning or how we might develop the kinds of curriculum modes that support problem‐based learning, and even what might count as a curriculum. This article seeks to unpack two of those issues by drawing on research into facilitation and modes of curriculum practice.
The origins of molecular biology: A Pedagogical tool for the professional development of pre‐college science teachers
Authors: Silverman Philip M.
Published: February 2007

We examine the science and pedagogy behind a historical approach to the professional development of pre‐college science (primarily biology) teachers. Our intention is to reach professional scientists, who, as a group, are uniquely qualified to provide experience and insights essential to this approach. The underlying research for this article has been carried out during an 8‐week summer course for selected rural high school science teachers at the Oklahoma Medical Research Foundation in Oklahoma City, Oklahoma. Now configured as part of the broader Oklahoma Science Project, the Summer Course has been held during each of the past 10 years. The result of our experiences is a learning model that incorporates a research dynamic based on peer interactions and Robert K. Merton's concept of specified ignorance. This dynamic is organized around carefully chosen research topics based on simple, quantitative assays that played important roles in the development of our current understanding of genetic chemistry. These assays are starting points for rationally conceived research projects developed by the teachers themselves. We illustrate this approach with the plaque assay for bacterial viruses.
Initiation of protein biosynthesis in eukaryotes
Authors: Merrick William C.
Published: February 2007

Initiation of protein synthesis in eukaryotes is important both mechanically, because it selects the translation reading frame, and biologically, because it is the primary site for regulation of translation. For approximately 95–98% of the cellular mRNAs, formation of an initiation complex follows a rather specific pathway that is broken down into seven discrete steps. Although almost 35 peptides in 12–14 translation initiation factors participate in this process, three appear to have dominant roles: eukaryotic initiation factor (eIF)3, in building a pool of 40S subunits; eIF2, in binding the initiator tRNA (tRNAi) to the 40S subunit; and eIF4F in activating the mRNA and binding it to the 40S subunit. This process requires both ATP and GTP. The resulting 80S initiation complex contains both the tRNAi and the mRNA, with the anticodon of the tRNAi correctly base paired with the initiating AUG code word. Regulation of translation focuses mostly on controlling the activity of either eIF2 or eIF4F and this regulation has different consequences. Reduction in eIF2 activity influences all mRNA approximately the same, whereas reduction in eIF4F activity drives competition between mRNAs.
Teaching molecular biology to undergraduate biology students: An illustration of protein expression and purification*
Authors: Sommer César Adolfo , Silva Flávio Henrique , Novo Maria Teresa Marques
Published: February 2007

Practical classes on protein expression and purification were given to undergraduate biology students enrolled in the elective course “Introduction to Genetic Engineering.” The heterologous expression of the green fluorescent protein (GFP)* of Aequorea victoria is an interesting system for didactic purposes because it can be viewed easily during experiments. The students were provided with basic information about the molecular features and applications of the GFP in molecular biology, the available heterologous expression systems, and the theoretical and experimental details of GFP expression in Escherichia coli and its purification. E. coli BL21‐competent cells were transformed with the pET28a expression vector containing the GFP gene fused to a histidine (His) tag. During the induction of a transformed clone by isopropylthiogalactoside, a time course for GFP expression was analyzed by SDS‐PAGE, and the expression was also visualized by the increasing green fluorescence of the bacterial culture. After cellular disruption, protein purification was illustrated by affinity chromatography of the His‐tagged protein in a nickel column. Eluted fractions containing imidazole in increasing concentrations were analyzed visually and also by SDS‐PAGE, demonstrating the role of imidazole in protein recovery by competition with nonspecific proteins and the His‐tagged protein. The results obtained and the experimental factors involved in protein expression, solubilization, and folding were discussed following the laboratory experiments. These practical classes allowed several current approaches to molecular biology to be demonstrated rapidly and helped underscore some of the topics taught during the course.
A response to BIO 2010: Transforming undergraduate education for future research biologists, from the perspective of the biochemistry and molecular biology major program at Kenyon College
Authors: Slonczewski Joan L. , Marusak Rosemary
Published: February 2007

The National Research Council completed a major study of undergraduate biology education, BIO 2010‐Transforming Undergraduate Education For Future Research Biologists (BIO 2010), funded by the Howard Hughes Medical Institute and the National Institutes of Health. The BIO 2010 report recommends that biology pedagogy should use an interdisciplinary approach incorporating a strong basis in mathematics and physical sciences. Many of the aims of BIO 2010 can be met by an interdisciplinary major program such as that of Biochemistry and Molecular Biology at Kenyon College. The Kenyon program effectively encourages students to connect biology with chemistry and mathematics and to develop a sound basis for research in the biological sciences. A continuing challenge is to balance the needs for depth of physical and mathematical understanding and breadth of diversity in biology.
Implementing the recommended curriculum in biochemistry and molecular biology at a regional comprehensive university through a biology/chemistry double major: The minnesota state university moorhead experience*
Authors: Wallert Mark , Brisch Ellen , Chastain Chris , Malott Michelle , Provost Joseph
Published: February 2007

Minnesota State University Moorhead (MSUM) is a regional comprehensive university that is part of the Minnesota State Colleges and Universities (MnSCU) system. The current student population consists of ∼7,600 full‐ and part‐time students who are enrolled in one of 135 majors that lead to baccalaureate degrees. MSUM is committed to excellence in science teaching and research for undergraduates. It is an institutional member of the Council on Undergraduate Research and has three faculty members participating in Project Kaleidoscope (PKAL) Faculty for the 21st Century. Fourteen years ago, MSUM renewed its effort to have faculty participate in active research. All science faculty members hired since that time have been required to establish research programs. The primary purpose for the faculty engaging in ongoing research projects is to involve undergraduates in a meaningful research experience, thus training these students to become scientists.
A colorimetric process to visualize erythrocyte exovesicles aggregates*
Authors: Saldanha Carlota , Santos Nuno C. , Martins‐Silva J.
Published: February 2007

A biochemistry laboratory class protocol is described in order to create an opportunity for students to apply by doing the theoretical concepts underlying biomolecules and vesicles properties, together with the principles of centrifugation and colorimetric methodologies. Through simple procedures the students will i) observe the segregation of the vesicles suspensions into two separate phases (phtalate esters gradient and vesicle aggregates); ii) visualize the vesicle aggregates by protein, enzyme, and phospholipids coloration processes; and iii) discuss and explain the visualized colors by proper biochemical reactions. The aims, objectives, methodology of teaching/learning, and assessment for this laboratory class are indicated.
An introductory undergraduate course covering animal cell culture techniques
Authors: Mozdziak Paul E. , Petitte James N. , Carson Susan D.
Published: February 2007

Animal cell culture is a core laboratory technique in many molecular biology, developmental biology, and biotechnology laboratories. Cell culture is a relatively old technique that has been sparingly taught at the undergraduate level. The traditional methodology for acquiring cell culture training has been through trial and error, instruction when undertaking the first graduate student position, or instruction when hired for a specific industrial cell culture position. However, cell culture is an important candidate course for any biotechnology‐related training program because it is a technique that must be performed by investigators before they perform many molecular procedures, and vertebrate cell culture is becoming increasingly important for biomanufacturing of therapeutic proteins. Therefore, a cell culture techniques course is an important offering for undergraduate students who aspire to graduate training, and also undergraduate students who will seek employment with biotechnology companies immediately after graduation. Recently, a cell culture techniques course was developed and delivered to students at North Carolina State University as a component of an undergraduate Biotechnology minor curricula. Currently, the instructors at North Carolina State University are seeking to provide students with the necessary technical and critical reasoning skills to successfully perform animal cell culture.
Diphenyl carbazide restores electron transport in isolated, illuminated chloroplasts after electron transport from water has been eliminated by mild heat treatment
Authors: Dean Rob L. , Pocock Tessa
Published: February 2007

Freshly isolated, illuminated chloroplasts oxidize water and transfer the resulting electrons through the photosynthetic electron transport chains in their thylakoid membranes to the artificial electron acceptor, dichlorophenol indophenol (DCPIP). As a consequence, DCPIP is reduced and the decline in absorbance over time can be used to measure the rate of electron transfer. When gently heated, chloroplasts lose the capacity to oxidize water and the transfer of electrons to DCPIP is eliminated. Electron transport through chloroplasts to DCPIP is restored in the presence of the artificial electron donor diphenylcarbazide (DPC). If students gain experience with the DCPIP photoreduction assay and are given information on normal chloroplast function, they should be able to predict the behavior of heat‐treated chloroplasts in a variety of experimental conditions. A number of such predictions are outlined and tested. The experiments can all be conducted with a limited repertoire of equipment and easily prepared solutions. Consequently, this work is well suited to an investigative study in which each student group, in consultation with instructors, can make and test its own prediction. The ways in which changing different variables can affect the quality of the experimental results is emphasized. Additional studies, on measurements of rates of oxygen evolution and emitted chlorophyll fluorescence, are briefly described to support the inferences that heat‐treated chloroplasts do not oxidize water and that the vectorial transfer of electrons through them to DCPIP is identical to that in untreated chloroplasts.
Estrogen regulation of the expression of a cytochrome P450 isoenzyme*
Authors: Szeberenyi Jozsef
Published: February 2007

Terms to be familiar with before you start to solve the test: cytochrome P450 enzymes, xenobiotics, estrogens, steroid receptors, gene induction, SDS, chromatin, immunoprecipitation, immunoglobulin G (IgG), agarose gel electrophoresis, promoter, transcription factors, enhancer, histones, nonhistone proteins, PCR, Taq polymerase, primers, oligonucleotides, radioactive labeling, nondenaturing PAGE.
From gene to protein: Prostatic acid phosphatase: Structure and expression of gene and protein*
Authors: Laidler Piotr , Kuciel Radoslawa , Dulińska Joanna , Gil Dorota , Mazurkiewicz Aleksandra , Wróbel Maria
Published: February 2007

A set of classes for medical students is designed to reinforce an understanding of the basic laboratory methods of molecular biology and protein biochemistry in the context of a clinically important problem, prostate gland pathology. Students examine the gene coding for prostatic acid phosphatase and they assay expression of the gene in different lines of prostate cancer cell cultures (LNCaP and PC‐3). The three‐dimensional structure of the expressed protein is also investigated, in relation to its catalytic function. Students are encouraged to collect data for their experiments and to perform laboratory exercises on their own. The theory and practice should stimulate the students' discussion of various fields of biochemistry and molecular biology.
A complete approach for recombinant protein expression training: From gene cloning to assessment of protein functionality*
Authors: Novo M. Teresa Marques , Soares‐Costa Andrea , de Souza Antonia Q. L. , Figueira Ana Carolina M. , Molina Gustavo C. , Palacios Carlos A. , Kull Claudia R. , Monteiro Izabel F. , Baldan‐Pineda Paulo H. , Henrique‐Silva Flavio
Published: February 2007

A practical course was given to undergraduate biology students enrolled in the elective course “Introduction to Genetic Engineering” at the Federal University of São Carlos (UFSCar), São Paulo, Brazil. The goal of the course was to teach current molecular biology tools applied to a real research situation that could be reported by the students themselves. The purpose was to produce a plant recombinant protein and demonstrate a heretofore unreported biological activity. Cystatins, natural inhibitors of cysteine proteases, were proposed for these studies. Initially, the students searched for plant cystatin cDNA sequences in the NCBI databases and selected the Oryzacystatin I gene (ocI) from rice, Oriza sativa, as the target gene for this study. Total RNA was extracted from rice‐germinating seeds and primers containing restriction sites for NdeI and EcoRI were designed based on the ocI cDNA sequence and then used to amplify the open reading frame (ORF). RT‐PCR amplification provided a band of the expected size for ocI ORF (309 bp). The PCR product was cut with NdeI and EcoRI restriction enzymes and cloned directly in the pET28a expression vector digested with the same enzymes. A pET28‐ocI recombinant clone was selected, checked by sequencing, and used to transform Escherichia coli BL21 (DE3) expression strain. After induction of the bacteria with isopropylthiogalactoside and cellular disruption, the His‐tagged OCI protein, present mainly in the soluble fraction, was purified by affinity chromatography in a nickel column. The purified protein was successfully used to inhibit fungal growth (Trichoderma reesei). The results were discussed extensively and the students contributed to the writing of this article, of which they are co‐authors.
Bioinformatics training programs are hot but the labor market is not*
Authors: Black Grant C. , Stephan Paula E.
Published: February 2007

Based on a survey of academic training programs and an analysis of advertised job openings, we conclude that the labor market in bioinformatics has changed dramatically from the 1990s to the early 2000s. The number of academic training programs, as well as enrollment in these programs, expanded rapidly during this period. The expansion has created a sizeable pipeline of students who will matriculate from these programs in the near future. Yet, at the same time that this expansion in training programs occurred, demand in the bioinformatics market declined and its origins have shifted largely from industry to academe. Unless conditions in industry change dramatically in the next few years, it is likely that trainees from these programs will have difficulty finding the expected jobs in industry.
A guided inquiry experiment for the measurement of activation energies in the biophysical chemistry laboratory: Decarboxylation of pyrrole‐2‐carboxylate
Authors: Hutchinson Kelly M. , Bretz Stacey Lowery , Mettee Howard D. , Smiley Jeffrey A.
Published: February 2007

A laboratory experiment for undergraduate biophysical chemistry is described, in which the acid concentration and temperature dependences of the decarboxylation of pyrrole‐2‐carboxylate are measured using a continuous ultraviolet (UV) spectrophotometric assay. Data collection and analysis are structured using principles of guided inquiry. Data leading to the calculation of multiple rate constants at varying temperatures and acid concentrations can be collected within one laboratory period, using inexpensive reagents and standard instrumentation. These experiments permit determination of activation energies that are lower at high acid concentration, indicative of a subtle change in the reaction mechanism with decreasing pH. The reaction is readily observable by students as they collect UV spectrophotometry data, and the decarboxylation reaction is related to biologically relevant enzymatic reactions.
From problem‐based learning to interrupted lecture: Using case‐based teaching in different class formats
Authors: Hodges Linda C.
Published: February 2007

Although many examples of using problem‐based learning and case studies in teaching science occur in the literature, instructors may still have questions about when, why, and how to use these strategies, especially in different class settings. In this article, I describe my experience in using these pedagogical methods in three different class formats: a small biochemistry survey course, which I taught entirely as problem‐based learning; a small organic chemistry class in which I used some case‐based teaching with students working collaboratively in groups; and a large lecture‐style biochemistry survey course in which I used cases as homework and in‐class exercises. In each instance I describe the apparent benefits and drawbacks of each approach, drawing on the literature of how these methods may best promote learning. As part of this discussion I pose questions for instructors to help them decide which form of case‐based teaching best fits with their particular teaching goals and class contexts.
Analysis of DNA replication in plant cells*
Authors: Szeberenyi Jozsef
Published: February 2007

Terms to be familiar with before you start to solve the test: cell division, phases of the cell cycle, phases of mitosis, chromosomes, chromatids, [3H]thymidine, pulse labeling, autoradiography.
Often ignored facts about the control of the 2‐oxoglutarate dehydrogenase complex
Authors: Strumilo Slawomir
Published: February 2007

Information about the control of the activity of the 2‐oxoglutarate dehydrogenase complex (OGDHC), a key enzyme in the citric acid cycle, is not well covered in the biochemical education literature, especially as it concerns the allosteric regulation of OGDHC by adenine nucleotide and ortophosphate. From experimental work published during the last 25 years, the following basic view is clear: (a) animal OGDHC is very sensitive to ADP, Pi, and Ca2+; (b) these positive effectors increase the affinity of OGDHC to 2‐oxoglutarate; (c) OGDHC is inhibited by ATP, NADH, and succinyl‐CoA; (d) the ATP effect is realized mainly via opposition to ADP activation; (e) NADH, in addition to inhibiting the dihydrolipoamide dehydrogenase component of the enzyme complex (competitively versus NAD+) decreases the affinity of 2‐oxoglutarate dehydrogenase to its substrate; (f) bacterial and plant OGDHC are activated by AMP instead of ADP. These main effects form the basis of short term regulation of OGDHC. It is desirable that such information should reach biochemistry students.
SigrafW: An easy‐to‐use program for fitting enzyme kinetic data
Authors: Leone Francisco Assis , Baranauskas José Augusto , Furriel Rosa Prazeres Melo , Borin Ivana Aparecida
Published: February 2007

SigrafW is Windows‐compatible software developed using the Microsoft® Visual Basic Studio program that uses the simplified Hill equation for fitting kinetic data from allosteric and Michaelian enzymes. SigrafW uses a modified Fibonacci search to calculate maximal velocity (V), the Hill coefficient (n), and the enzyme‐substrate apparent dissociation constant (K). The estimation of V, K, and the sum of the squares of residuals is performed using a Wilkinson nonlinear regression at any Hill coefficient (n). In contrast to many currently available kinetic analysis programs, SigrafW shows several advantages for the determination of kinetic parameters of both hyperbolic and nonhyperbolic saturation curves. No initial estimates of the kinetic parameters are required, a measure of the goodness‐of‐the‐fit for each calculation performed is provided, the nonlinear regression used for calculations eliminates the statistical bias inherent in linear transformations, and the software can be used for enzyme kinetic simulations either for educational or research purposes.

Persons interested in receiving a free copy of the software should contact Dr. F. A. Leone.
The mitochondrial phosphate‐to‐oxygen ratio is not an integer
Authors: Silverstein Todd
Published: February 2007

For many years, mitochondrial phosphate‐to‐oxygen ratios were believed to be integral values: 3 for ATP/NADH and 2 for ATP/FADH2. The chemiosmotic theory has demonstrated that this need not be the case because coupling sites do not directly synthesize ATP. Coupling between coupling site redox reactions and ATP synthesis occurs via proton pumping and proton flow, respectively. Most recent measurements support phosphate‐to‐oxygen ratios of ∼2.5 ATP/NADH and 1.5 ATP/FADH2; these values match those expected from chemiosmotic calculations. Faculty members who teach metabolism are encouraged to reflect these developments in their lecture notes.
One hundred years of American Women in biochemistry *
Authors: Wolfson Adele J.
Published: February 2007

The occasion of ASBMB's 100th anniversary provides an opportunity to consider the history and accomplishments of women biochemists over the past century. American women in biochemistry have ranged from the highly visible, such as Nobel Prize winners Gert Cori and Gertrude Elion, to those who contributed in relative obscurity to equally dramatic breakthroughs. Women with Ph.D.s in biochemistry established departments of public health, nutrition, and home economics when more prestigious academic departments were not open to them. As in all fields of science, women have made great progress in biochemistry in recent years, and ASBMB reflects that progress; the past, current, and incoming presidents of the Society are all women. The data and biographical information presented here should provide a basis for inclusion of more women in recounting the history of biochemistry.
Thoughts on the teaching of metabolism
Authors: Metzger Robert P.
Published: February 2007

Systems biology, metabolomics, metabolic engineering, and other recent developments in biochemistry suggest that future biochemists will require a detailed familiarity with the compounds and pathways of intermediary metabolism and their biochemical control. The challenge to the biochemistry instructor is the presentation of metabolic pathways in a manner that allows student creativity in learning the pathways and their components. One approach that does permit the use of problem solving for the study of metabolic pathways involves following the fate of 13C, 14C, or 15N labels, presented originally in the structure of an important metabolic starting compound, through relevant metabolic pathways. This method allows the presentation and study of problems in which such an isotopic label can be traced through two or more metabolic pathways, thus illustrating how these pathways are interconnected. The understanding that all the pathways of intermediary metabolism are interconnected provides opportunities to discuss their metabolic control by such mechanisms as signaling, feedback inhibition, location in organelles, coenzyme levels, and coenzyme recycling rates. The method is illustrated by following the fate of 14C labels through anaerobic glycolysis, gluconeogenesis, and fatty acid transport, β‐oxidation, and ketone body formation. Cholesterol biosynthesis and heme formation are used to show that presentations of long and complex pathways can demonstrate important biochemical concepts by following the fate of an isotopic label using only the most important intermediates. Problems based on tracing radioactive labels through one or more metabolic pathways allow the use of cooperative learning techniques.
An exercise to estimate differential gene expression in human cells
Authors: Chaudhry M. Ahmad
Published: February 2007

The expression of genes in cells of various tissue types varies considerably and is correlated with the function of a particular organ. The pattern of gene expression changes in diseased tissues, in response to therapy or infection and exposure to environmental mutagens, chemicals, ultraviolet light, and ionizing radiation. To better understand the disease at the molecular level, the alterations in gene expression can provide clues to identify the cellular pathways involved. There are a wide variety of techniques that have been developed to study differential gene expression. Most of these methodologies are technically challenging. We developed an exercise to examine and quantify differential gene expression based on reverse transcription‐PCR. Our goal was to have this technique successfully performed by college level students. This technique is easy to perform and does not require the use of radioactive substances. We examined the ionizing radiation‐induced changes in gene expression in human cells. After exposure to ionizing radiation, total RNA was isolated at 4, 8, 12, and 24 h. The RNA was converted to cDNA and subjected to PCR amplification using gene‐specific primers and commercially available β‐actin internal standards in a multiplex format. The differences in gene expression were quantified with freely available imaging software. This technique should have a wide application to investigate differential gene expression in a variety of organisms and under various experimental and treatment conditions.
Using advanced problems in introductory courses: Some sample problems and why they work
Authors: Mowshowitz Deborah
Published: February 2007

This article presents several typical problems used in an introductory course in molecular biology and discusses why the problems are effective at increasing learning.
Using yeast to easily determine mitochondrial functionality with 1‐(4,5‐dimethylthiazol‐2‐yl)‐3,5‐diphenyltetrazolium bromide (MTT) assay
Authors: Sánchez Norma Silvia , Königsberg Mina
Published: February 2007

Mitochondrial respiration is an important topic in biochemical courses, but at the same time it is a very arduous and challenging subject for the students to comprehend. In teaching laboratories, experiments are also difficult to perform because mitochondrial isolation as well as oxygen consumption determinations require expensive equipment, rarely present in Mexican teaching laboratories. For this purpose, we adapted the 1‐(4,5‐dimethylthiazol‐2‐yl)‐3,5‐diphenyltetrazolium bromide (MTT) assay, usually done with cell lines, to corroborate mitochondrial respiration in yeast. We used the reduction of the MTT to learn and distinguish the respiratory yield between different substrates as well as the effects of respiratory inhibitors. This is an economical, easily done laboratory experiment that quantifies mitochondrial functionality and provides a good idea of the respiratory process.
“Mini‐array” transcriptional analysis of the Listeria monocytogenes lecithinase operon as a class project: A student investigative molecular biology laboratory experience*
Authors: Christensen Douglas , Jovic Marko
Published: February 2007

This report describes a molecular biotechnology‐based laboratory curriculum developed to accompany an undergraduate genetics course. During the course of a semester, students researched the pathogen, developed a research question, designed experiments, and performed transcriptional analysis of a set of genes that confer virulence to the food‐borne pathogen, Listeria monocytogenes. Gene fragments were amplified via PCR and utilized in “mini‐arrays,” a dot‐blot‐based format suitable for the simultaneous transcriptional analysis of multiple genes. The project provides exposure to a wide range of molecular techniques and can be easily modified for variations in class size. Data are generated at various steps of the process, allowing for student interpretation, troubleshooting, and assessment opportunities.
Fluorescence analysis of sulfonamide binding to carbonic anhydrase
Authors: Wang Sheila C. , Zamble Deborah B.
Published: February 2007

A practical laboratory experiment is described that illustrates the application of fluorescence resonance energy transfer to the study of protein‐ligand binding. The affinities of wild‐type and mutant human carbonic anhydrase II for dansylamide were determined by monitoring the increase in ligand fluorescence that occurs due to energy transfer from tryptophan residues near the enzyme active site. In a subsequent experiment, the binding constant of azetazolamide, a weaker fluorophore but a stronger ligand, is measured by competition with dansylamide. This simple experiment introduces students to the widely used technique of fluorescence spectroscopy and to the determination of protein‐ligand binding constants.
Helping parents to manage infant crying and sleeping: a review of the evidence and its implications for services
Authors: James‐Roberts Ian St
Published: February 2007

Infant crying and night waking are common concerns for parents, costly problems for health services and may trigger infant abuse or lead to serious child disturbances. Parents are given contradictory advice on how to manage infant crying and sleeping, indicating the need for evidence‐based guidance. This review of recent research draws distinctions between infant crying and sleeping problems, between the problem identified by parents and the infant behaviour underlying the problem, between different types of crying behaviour and their causes, and between the types of cases which present at different ages. It proposes that the two main approaches to parenting advocated by baby‐care experts, ‘infant‐demand’ and ‘structured’ parenting, have different benefits, and costs. Comparative studies have found that infant‐demand parenting is associated with low amounts of fussing and crying in the first three months of age, but with night waking which continues beyond three months. Randomised controlled trials have provided evidence that structured parenting leads to more overall fussing and crying during the first three months, but reduced night waking and crying after that. The findings are translated into recommendations for preventing and treating infant crying and sleeping problems, for policy debate, and for further research. Copyright © 2007 John Wiley & Sons, Ltd.
Theoretical Evaluation of the Substrate‐Assisted Catalysis Mechanism for the Hydrolysis of Phosphate Monoester Dianions
Authors: Iché‐Tarrat Nathalie , Ruiz‐Lopez Manuel , Barthelat Jean‐Claude , Vigroux Alain
Published: February 2007

Quantum chemistry methods coupled with a continuum solvation model have been applied to evaluate the substrate‐assisted catalysis (SAC) mechanism recently proposed for the hydrolysis of phosphate monoester dianions. The SAC mechanism, in which a proton from the nucleophile is transferred to a nonbridging phosphoryl oxygen atom of the substrate prior to attack, has been proposed in opposition to the widely accepted mechanism of direct nucleophilic reaction. We have assessed the SAC proposal for the hydrolysis of three representative phosphate monoester dianions (2,4‐dinitrophenyl phosphate, phenyl phosphate, and methyl phosphate) by considering the reactivity of the hydroxide ion toward the phosphorus center of the corresponding singly protonated monoesters. The reliability of the calculations was verified by comparing the calculated and the observed values of the activation free energies for the analogous SN2(P) reactions of F− with the monoanion of the monoester 2,4‐dinitrophenyl phosphate and its diester analogue, methyl 2,4‐dinitrophenyl phosphate. It was found that the orientation of the phosphate hydrogen atom has important implications with regard to the nature of the transition state. Hard nucleophiles such as OH− and F− can attack the phosphorus atom of a singly protonated phosphate monoester only if the phosphate hydrogen atom is oriented toward the leaving‐group oxygen atom. As a result of this proton orientation, the SAC mechanism in solution is characterized by a small Brønsted coefficient value (βlg=−0.25). This mechanism is unlikely to apply to aryl phosphates, but becomes a likely possibility for alkyl phosphate esters. If oxyanionic nucleophiles of pKa<11 are involved, as in alkaline phosphatase, then the SN2(P) reaction may proceed with the phosphate hydrogen atom oriented toward the nucleophile. In this situation, a large negative value of βlg (−0.95) is predicted for the substrate‐assisted catalysis mechanism.
N‐Octylquinolinium Tribromide: A Task Specific Quinoline Based Ionic Liquid as a New Brominating Agent.
Authors: Kaushik M. P. , Polshettiwar Vivek
Published: February 2007

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract, please click on HTML or PDF.
Adenylate kinase in sea urchin embryonic cilia
Authors: Kinukawa Masashi , Vacquier Victor D.
Published: February 2007

Sea urchin embryos swim by ciliary movement. Hypertonic shock causes deciliation and loss of motility. Within 2–4 h, cilia regenerate and the embryos swim again. Regeneration of cilia occurs multiple times. The adenylate kinase (AK) activity of isolated cilia was studied. A 130‐kDa Sp‐AK isozyme, present in sperm flagella, is also present in embryonic cilia. AK activity is responsible for ∼93% of nonmitochondrial ATP regeneration from ADP in embryonic cilia. This is unlike sea urchin sperm flagella, where ∼31% of the nonmitochondrial ATP regeneration is from the 130‐kDa Sp‐AK isozyme and ∼69% from the flagellar creatine kinase (Sp‐CK). Embryos were deciliated 1–3 times and after a 2‐h period of regeneration the major ciliary axonemal proteins such as the tubulins appeared constant in amount. However, a moderate decrease in ATPase activity, and a large decrease of total AK activity, were measured. The decrease in AK activity paralleled the decrease in embryo swimming velocity. Embryos were deciliated once and cilia regeneration followed for 4 h. ATPase activity recovered to control levels by 3 h, but AK activity and swimming velocity remained lower than in controls. Detergent solubility data and kinetic experiments indicate that, in addition to the 130‐kDa Sp‐AK, there is at least one additional AK isozyme in embryonic cilia. Analysis of the S. purpuratus genome indicates five AK isozymes in addition to the 130‐kDa Sp‐AK isozyme. Decreased swimming velocity of embryos with regenerated cilia suggests that regenerated cilia are not as functionally perfect as naturally grown cilia. Cell Motil. Cytoskeleton 2007. © 2007 Wiley‐Liss, Inc.
Effects of cross‐fostering on cocaine‐induced conditioned taste aversions in Fischer and Lewis rats
Authors: Roma Peter G. , Davis Catherine M. , Riley Anthony L.
Published: February 2007

The systematic comparison between Fischer and Lewis rats is a popular animal model of genetic factors in drug abuse. Although genetic and environmental factors interact to affect drug abuse in humans, analogous effects have not yet been reported within the Fischer–Lewis model. In order to assess the contributions and interaction of genotype and early maternal environment on responses to a drug of abuse, the present study employed a cross‐fostering design, where male and female Fischer and Lewis pups were reared by unrelated dams of their own strain (in‐fostered) or of the other strain (cross‐fostered). As adults, rats from both strains were tested for their ability to acquire a conditioned taste aversion to a novel saccharin solution that had been repeatedly paired with an injection of cocaine (32 mg/kg, subcutaneous). In‐fostered Fischer females acquired significantly weaker aversions than in‐fostered Lewis females across the multiple saccharin‐cocaine pairings. However, cross‐fostered Fischer females exhibited aversions that were not only significantly stronger than their in‐fostered Fischer counterparts, but identical to all groups of the Lewis genotype. No strain differences or cross‐fostering effects were observed in the males. The data with the female subjects cannot be accounted for simply by the genetic strain of the subjects and demonstrate a clear gene‐environment interaction effect on responses to the aversive effects of cocaine in Fischer and Lewis rats. Implications for studying maternal behavior as a source of epigenetic modulation of drug abuse vulnerability were discussed. © 2007 Wiley Periodicals, Inc. Dev Psychobiol 49: 172–179, 2007.
Children's skin conductance level and reactivity: Are these measures stable over time and across tasks?
Authors: El‐Sheikh Mona
Published: February 2007

The stability of children's skin conductance level during baselines (SCL‐B) and SCL reactivity (SCL‐R) were examined longitudinally. During two laboratory sessions (T1 and T2), 2 years apart, children participated in procedures during which they were exposed to two stressors namely exposure to an audiotaped conflict between two adults, and a problem solving task. Children ranged in age between 6 and 13 years at T1. Measures of SCL‐B and SCL‐R during the two stressors were obtained. Findings illustrated the temporal stability of SCL‐B and SCL‐R to the star‐tracing task over 2 years. Results also indicated stability in SCL‐R to the two stressors (argument and problem‐solving) examined within the same session at either T1 or T2. These results support the proposition that SCL‐B and SCL‐R constitute stable individual differences at the ages examined, and build on the scant longitudinal literature on psychophysiological development in children. © 2007 Wiley Periodicals, Inc. Dev Psychobiol 49: 180–186, 2007.
Cyanido‐Bridged Dimetallic Complexes Derived from Manganese(III) Schiff Bases and Pentacyanidonitrosylchromate(I): Synthesis, Crystal Structure and Magnetic Properties
Authors: Ni Zhong‐Hai , Zheng Lei , Zhang Li‐Fang , Cui Ai‐Li , Ni Wei‐Wei , Zhao Chong‐Chao , Kou Hui‐Zhong
Published: February 2007

Four cyanido‐bridged MnIII–CrI heterodimetallic complexes have been synthesized by the reactions of MnIII Schiff‐base complexes with the [Cr(CN)5(NO)]3– building block. X‐ray diffraction analysis reveals that [Mn(5‐Cl‐salen)(H2O)2][Mn2(5‐Cl‐salen)2(H2O)1.5Cr(CN)5(NO)] (1) has a trinuclear structure, and [Mn3(salen)3(H2O)(CH3CN)Cr(CN)5(NO)]·0.5CH3CN·0.5MeOH·2H2O (2) and [Mn(5‐Br‐salen)(H2O)(CH3OH)][Mn2(5‐Br‐salen)2(CH3OH)Cr(CN)5(NO)]·2H2O (3) have novel 1D chain topological structures, whereas [{Mn(saltmen)}4Cr(CN)5(NO)]ClO4·3H2O (4) displays a 2D layered network structure. Magnetic measurements show that the magnetic interaction between the MnIII and CrI ions bridged by a cyanido group is antiferromagnetic for complexes 1–4. The magnetic fitting for the trinuclear complex 1 gives the magnetic coupling constant J = –10.4(1) cm–1 and J′ = –1.29(2) cm–1 based on the isotropic Hamiltonian Ĥ = –2JŜCr[ŜMn(1) + ŜMn(2)] – 2J′ŜMn(1)ŜMn(2). Complex 2 exhibits 3D antiferromagnetic ordering (TN = 3.5 K) with a metamagnetic character below TN. Complex 3 exhibits an obvious frequency‐dependent AC magnetic susceptibility below 4.0 K, which is due to the spin‐glass behavior. The layered complex 4 displays 3D ferromagnetic ordering with Tc = 6.4 K due to the ferromagnetic interaction between the ferrimagnetic layers. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)
A Two‐Dimensional, Hydrogen‐Bond‐Cross‐Linked Molybdenum(VI) Network Polymer with Catalytic Activity
Authors: Luan Yi , Wang Ge , Luck Rudy L. , Yang Mu
Published: February 2007

The new hybrid inorganic–organic polymer [MoO2Cl2(H2O)2]·(H2dipy‐pra)Cl2 (1), where dipy‐pra = 1,3‐bis(4‐pyridyl)propane, has been synthesized and crystallographically characterized. MoO2Cl2(H2O)2 and the [H2dipy‐pra]2+ cation are cross‐linked by Mo–H2O···Cl and H2dipy‐pra···Cl hydrogen bonds to form a two‐dimensional layer structure. Complex 1 is an efficient catalyst, with H2O2 as the oxygen‐source oxidant and NaHCO3 as the cocatalyst, in the epoxidation of olefinic compounds under ambient conditions.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)
Bioinspired Metal‐Catalysed Doebner–Knoevenagel Condensations between Malonic Acid Half Thioesters and Aldehydes
Authors: Berrué Fabrice , Antoniotti Sylvain , Thomas Olivier P. , Amade Philippe
Published: February 2007

Inspired by polyketide biosynthesis, unprecedented metal‐catalysed Doebner–Knoevenagel condensations between Malonic Acid Half Thioesters (MAHTs) and various aldehydes have been achieved through the use of Yb(OTf)3 as catalyst, mimicking a five‐domain polyketide synthase module for the first time. In the presence of 5‐methoxybenzimidazole, stereo‐ and regiocontrol of the reaction proved to be high. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)
NMR Investigation of the Bound Conformation of Natural and Synthetic Oligomannosides to Banana Lectin
Authors: Clavel Caroline , Canales Angeles , Gupta Garima , Cañada F. Javier , Penadés Soledad , Surolia Avadhesha , Jiménez‐Barbero Jesús
Published: February 2007

The conformational behaviour of three mannose‐containing oligosaccharides, namely, the α1→3[α1→6] trisaccharide, a heptasaccharide with α1→2, α1→3 and α1→6 linkages and a tetrasaccharide consisting of α1→3 and α1→2 linkages, when bound to banana lectin (BanLec) has been evaluated by trNOE NMR methods and docking calculations. It was found that the molecular recognition event involves a conformational selection process with only one of the conformations present in the free state of the sugar being recognised at the lectin binding site. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)