Intensive research and development of electrophoresis methodology and instrumentation during past decades has resulted in unique methods widely implemented in bioanalysis. While two‐dimensional electrophoresis and denaturing polyacrylamide gel electrophoresis in sodium dodecylsulfate are still the most frequently used electrophoretic methods applied to analyses of proteins, new miniaturized capillary and microfluidic versions of electromigrational methods have been developed. High‐throughput electrophoretic instruments with hundreds of capillaries for parallel separations and laser‐induced fluorescence detection of labeled DNA strands have been of (...) key importance for the scientific and commercial success of the Human Genome Project. Another powerful method, capillary isoelectric focusing with pressurized and pH‐driven mobilization, provides efficient separations and on‐line sensitive detection of proteins, bacteria and viruses. Electrophoretic microfluidic devices can integrate single‐cell injection, cell lysis, separation of its components and fluorescence or mass spectrometry detection. These miniaturized devices also proved the capability of single‐molecule detection. (shrink)

Preparative and analytical methods developed by separation scientists have played an important role in the history of molecular biology. One such early method is gel electrophoresis, a technique that uses various types of gel as its supporting medium to separate charged molecules based on size and other properties. Historians of science, however, have only recently begun to pay closer attention to this material epistemological dimension of biomolecular science. This paper substantiates the historiographical thread that explores the relationship between modern (...) laboratory practice and the production of scientific knowledge. It traces the historical development of gel electrophoresis from the mid-1940s to the mid-1960s, with careful attention to the interplay between technical developments and disciplinary shifts, especially the rise of molecular biology in this time-frame. Claiming that the early 1950s marked a decisive shift in the evolution of electrophoretic methods from moving boundary to zone electrophoresis, I reconstruct various trajectories in which scientists such as Oliver Smithies sought out the most desirable solid supporting medium for electrophoretic instrumentation. Biomolecular knowledge, I argue, emerged in part from this process of seeking the most appropriate supporting medium that allowed for discrete molecular separation and visualization. The early 1950s, therefore, marked not only an important turning point in the history of separation science, but also a transformative moment in the history of the life sciences as the growth of molecular biology depended in part on the epistemological access to the molecular realm available through these evolving technologies. (shrink)

Preparative and analytical methods developed by separation scientists have played an important role in the history of molecular biology. One such early method is gel electrophoresis, a technique that uses various types of gel as its supporting medium to separate charged molecules based on size and other properties. Historians of science, however, have only recently begun to pay closer attention to this material epistemological dimension of biomolecular science. This paper substantiates the historiographical thread that explores the relationship between modern (...) laboratory practice and the production of scientific knowledge. It traces the historical development of gel electrophoresis from the mid-1940s to the mid-1960s, with careful attention to the interplay between technical developments and disciplinary shifts, especially the rise of molecular biology in this time-frame. Claiming that the early 1950s marked a decisive shift in the evolution of electrophoretic methods from moving boundary to zone electrophoresis, I reconstruct various trajectories in which scientists such as Oliver Smithies sought out the most desirable solid supporting medium for electrophoretic instrumentation. Biomolecular knowledge, I argue, emerged in part from this process of seeking the most appropriate supporting medium that allowed for discrete molecular separation and visualization. The early 1950s, therefore, marked not only an important turning point in the history of separation science, but also a transformative moment in the history of the life sciences as the growth of molecular biology depended in part on the epistemological access to the molecular realm available through these evolving technologies. (shrink)

For the last ten years, within molecular life sciences, the reproducibility crisis discourse has been embodied as a crisis of trust in scientific images. Beyond the contentious perception of “questionable research practices” associated with a digital turn in the production of images, this paper highlights the transformations of gel electrophoresis as a family of experimental techniques. Our aim is to analyze the evolving epistemic status of generated images and its connection with a crisis of trust in images within that (...) field.From the 1980s to the 2000s, we identify two key innovations (precast gels and gel docs) leading to a “two‐tiered” gel electrophoresis with different standardization procedures, different epistemic statuses of the produced images and different ways of generating (dis)trust in images. The first tier, exemplified by differential gel electrophoresis (DIGE), is characterized by specialized devices processing images as quantitative data. The second tier, exemplified by polyacrylamide gel electrophoresis (PAGE), is described as a routine technique making use of image as qualitative “virtual witnessing.” The difference between these two tiers is particularly apparent in the ways images are processed, even though both tiers involve image digitization. Our account thus highlights different views on reproducibility within the two tiers. Comparability of images is insisted upon in the first tier while traceability is expected in the second tier. It is striking that these differences occur not only within the same scientific field, but even within the same family of experimental techniques. In the second tier, digitization entails distrust, whereas it implies a collective sentiment of trust in the first tier. (shrink)

During replication, the topology of DNA changes continuously in response to well‐known activities of DNA helicases, polymerases, and topoisomerases. However, replisomes do not always progress at a constant speed and can slow‐down and even stall at precise sites. The way these changes in the rate of replisome progression affect DNA topology is not yet well understood. The interplay of DNA topology and replication in several cases where progression of replication forks reacts differently to changes in DNA topology ahead is discussed (...) here. It is proposed, there are at least two types of replication fork barriers: those that behave also as topological barriers and those that do not. Two‐Dimensional (2D) agarose gel electrophoresis is the method of choice to distinguish between these two different types of replication fork barriers. (shrink)

A crucial part of the knowledge of molecular biologists is procedural knowledge, that is, knowledge of how to do things in laboratories. Procedural knowledge of molecular biologists involves both perceptual-motor skills and cognitive skills. We discuss such skills required in performing the most commonly used molecular biology techniques, namely, Polymerase Chain Reaction and gel electrophoresis. We argue that procedural knowledge involved in performing these techniques is more than just knowing their protocols. Creative exploration and experience are essential for the (...) acquisition of procedural knowledge in molecular biology. With enough experience, molecular biologists make intuitive judgments without recourse to analytical reasoning. We propose that procedural knowledge is intuitive recognition of the patterns of one's environment that are the most relevant for making a decision or acting appropriately. Finally, we argue that knowledge of molecular biologists requires an integration of procedural knowledge and propositional knowledge. (shrink)

Berichte zur Wissenschaftsgeschichte, Volume 45, Issue 3, Page 332-343, September 2022.

Berichte zur Wissenschaftsgeschichte, Volume 45, Issue 3, Page 332-343, September 2022.

It has long been thought that gene expression is tightly regulated in multicellular eukaryotes, so that expression profiles match functional profiles. This conception emerged from the assumption that gene activity is synonymous with gene function. This paradigm was first challenged by comparative protein electrophoresis studies showing extensive differences in expression patterns among related species. The paradigm is now being challenged by evolutionary transcriptomics using microarray technologies. Most gene expression profiles display features that lack any obvious functional significance. The so‐called (...) “ectopic” expression refers to the expression of genes at times and locations where the target gene is not known to have a function. However, ectopic expression might be associated with genuine function even if this function is not essential or has yet to be ascertained. Alternatively, ectopic expression might come about as a superfluous by‐product of regulatory systems, which would call for a revision of prevailing ideas about the specificity of gene regulation. We herein review available evidence for ectopic expression and the hypotheses proposed for its origin and evolution. We propose that ectopic expression must be regarded as part of an integrated phenotypic whole. It seems likely that ectopic expression represents a leak in the evolution of regulatory systems, but one that is endowed with considerable evolutionary possibilities. BioEssays 27:592–601, 2005. © 2005 Wiley Periodicals, Inc. (shrink)

Until quite recently, recombinant DNA technology was not able to deal with DNA molecules larger than 20–40 kb. This is a serious limitation for the study of mammalian, and in particular human genomes whose total length is approx. 3 × 106 kb, since the best resolution of genetic and chromosomal analysis is usually the rough equivalent of 1000–5000 kb. Three recently developed methods promise to bridge this gap: pulsed field gel electrophoresis, which can analyze megabase‐sized DNA fragments; cloning in (...) yeast, which can clone and propagate DNA fragments of several hundred kb; and jumping libraries, which allow ‘jumping’ over large distances along the chromosome. This review presents the current status of these very promising technologies. (shrink)

The replicon hypothesis, first proposed in 1963 by Jacob and Brenner(1), states that DNA replication is controlled at sites called origins. Replication origins have been well studied in prokaryotes. However, the study of eukaryotic chromosomal origins has lagged behind, because until recently there has been no method for reliably determining the identity and location of origins from eukaryotic chromosomes. Here, we review a technique we developed with the yeast Saccharomyces cerevisiae that allows both the mapping of replication origins and an (...) assessment of their activity. Two‐dimensional agarose gel electrophoresis and Southern hybridization with total genomic DNA are used to determine whether a particular restriction fragment acquires the branched structure diagnostic of replication initiation. The technique has been used to localize origins in yeast chromosomes and assess their initiation efficiency. In some cases, origin activation is dependent upon the surrounding context. The technique is also being applied to a variety of eukaryotic organisms. (shrink)

For many years, there has been a gap in our capacity to study the structure and organization of chromosomal DNA molecules. The very small genomes of some viruses and bacteriophages (≤ 50,000 bp or 50 kb) are amenable to analysis by conventional gel electrophoresis, while the extremely large DNA molecules (> 100,000 kb) comprising the chromosomes of higher eukaryotes have been analysed under the light microscope, using a range of banding and in situ hybridization techniques. However, intact DNA molecules (...) with sizes between these two extremes have been largely inaccessible experimentally. This gap has recently been bridged with the development of two‐dimensional electrophoretic procedures that allow the separation and purification of chromosome‐sized DNA molecules ranging from ∼ 50 kb to several thousand kb.1–3 There are currently two variations of the technique in use: pulsed field gradient (PFG) gel electrophoresis1,2 and orthogonal‐field‐alteration gel electrophoresis (OFAGE).3 Both are based on a common principle, differing primarily in the geometry of the electrodes. Already, they have been employed to determine the approximate chromosome sizes and numbers for a variety of lower eukaryotes, including yeast and several protozoa.2–10 These ‘molecular karyotypes’ provide fundamental information about the genomic organization of each organism, and allow very rapid construction of linkage maps. Surprisingly, they have also revealed a remarkable plasticity in the genomes of several lower eukaryotes. (shrink)

The book illustrates how Darwin's theory has evolved, about the development of the biological world before Darwin, and great changes that took place with the incorporation of statistics, and after Darwin's death of genetics and mathematics. The formation of 'Modern Synthesis', protein electrophoresis, Discovery of DNA opened new avenues for the study of evolution.

Historians of molecular biology have paid significant attention to the role of scientific instruments and their relationship to the production of biological knowledge. For instance, Lily Kay has examined the history of electrophoresis, Boelie Elzen has analyzed the development of the ultracentrifuge as an enabling technology for molecular biology, and Nicolas Rasmussen has examined how molecular biology was transformed by the introduction of the electron microscope (Kay 1998, 1993; Elzen 1986; Rasmussen 1997). 1 Collectively, these historians have demonstrated how (...) instruments and other elements of the material culture of the laboratory have played a decisive role in determining the kind and quantity of .. (shrink)

Background: Increasing resistance of Pseudomonas aeruginosa to ciprofloxacin in ICU/burn units has created a problem in the treatment of infections caused by this microorganism. -/- Methods: Fifty P. aeruginosa strains were isolated from burn patients hospitalized in the Kerman Hospital during May 1999-April 2000 and were tested for in-vitro sensitivity to different antibiotics by disc diffusion breakpoint assay. The isolates were subjected to minimum inhibitory concentration (MIC) test by agar dilution method. Existence of the plasmids was also investigated in the (...) isolates. -/- Results: Thirty-four patients infected with ciprofloxacin strains showed MIC of 8 mg/ml [p<0.001]. Sixteen patients were infected with sensitive strains exhibiting MIC range of 0.0125-0.125 ± 0.033 mg/ml. The isolates were also also resistant to other antibiotics [p<0.001]. Plasmid isolation and agarose gel electrophoresis (0.7%) revealed three plasmid bands in strains 8 and 16, and one band in strain 35. -/- Conclusion: The emergence of ciprofloxacin resistance of P. aeruginosa in burn patients is alarming since this antibiotic has only recently been introduced onto the market in Iran. One important observation was that some isolates exhibited cross resistance to other antibiotics. Furthermore, some strains were carriers of plasmids which might have acted as the potential source of acquired resistance in the hospital setting. -/- . (shrink)

The human genome contains about 30,000 genes, each creating several transcripts per gene. Transcript structures and expression are studied by high‐throughput transcriptomic techniques using microarrays. Generally, transcripts are not directly operating molecules, but are translated into functional proteins, post‐translationally modified by proteolysis, glycosylation, phosphorylation, etc., sometimes with great functional impact. Proteins need to be analyzed by proteomic techniques, less suited for high‐throughput. Two‐dimensional polyacrylamide gel electrophoresis (2D‐PAGE), separating thousands of proteins has developed slowly over the past quarter of a (...) century. This technique is now quite reproducible and suitable for differential proteomics, comparing normal and diseased cells/tissues revealing differentially regulated proteins. 2D‐PAGE is combined with protein‐identification methods, currently mass spectrometry (MS), which has been significantly improved over the last decade. Other proteomic techniques studying protein–protein interactions are now either established or still being developed, such as peptide or protein arrays, phage display, and the yeast two‐hybrid system. The strengths and weaknesses of these techniques are discussed. BioEssays 26:901–915, 2004. © 2004 Wiley Periodicals, Inc. (shrink)

AMBIS is a complete identification system which includes (1) a highly reproducible electrophoresis unit; (2) a beta‐scanner with the ability to rapidly locate and measure beta particle emission data from a variety of isotopes and surfaces; and (3) an IBM computer with a massive data storage capacity for the emission data plus subsequent manipulation of that data. Hence it provides a rapid facility for (1) classification of all types of micro‐organisms, (2) examination of cells of multicellular plants and animals, (...) (3) routine determination of molecular weights of one or more proteins in a complex mixture, (4) quantification of single or mixed compounds in a band, (5) scanning and quantification of dot and other blots, immunodiffusion precipitates and probes, (6) direct counting of radioactive solutions from RIA, column fractions, etc., without the need of expensive scintillation fluids with their consequent hazards, (7) circumventing the need of much time‐consuming auto‐radiography, and (8) setting up databases plus exchange and transference of data between AMBIS systems in different locations via floppy disk, modem or other forms of rapid transmission. (shrink)

This paper comments on five articles in the special issue “Circulating Images in the Life Sciences.” It sees the papers as unified by two themes. The first is their attention to the processes of legitimation. The second is the embedding of the images in textual cultures, which changed over time from the mid‐nineteenth century to the very recent past, most notably with the recent advent of digital culture.

DNA is an important agent not only in chemistry and biology but also in technology and modern culture. A number of books approach the double helix from different angles. These perspectives include (1) the science of DNA and genetics; (2) genetic engineering; (3) the ethics of manipulating genetic material; and (4) DNA in culture and religion. Various views of DNAprovide insights into human nature beyond its molecular composition.