bluuebunny/crossref_metadata_2025_split huggingface dataset:
Intracellular staining of enteric neurones
Authors: Lees Gordon M 
Published: November 2023

Abstract
It is now widely accepted that the enteric nervous system (ENS) provides neuroscientists with many challenges, one of the most important problems being the identification of neuronal pathways involved in intestinal reflexes, particularly intrinsic reflex responses, such as peristalsis. The intrinsic neurones of the gastrointestinal tract lie in ganglionated plexuses between the (outer) longitudinal muscle and the circular muscle layers (myenteric plexus) and in the submucosa (inner and outer submucous plexuses) (Figure J). Morphologically, elcctro physiologically, immunohistochcmically, and pharmacologically these enteric neurones show an amazing diversity, both between and within the plexuses (1-10).
Making of the atlas
Authors: Singh Juswinder , Thornton Janet M 
Published: November 2023

Abstract
This section describes how the information in the atlas was derived. It describes the protein dataset used, the definition of the side chain interactions and their geometries, and the use of local coordinate systems to generate three-dimensional distributions of interacting pairs. The suite of programs used for the calculation and analysis of the interacting pairs is called SIRIUS (Singh and Thornton 1990), and a flowchart of the approach is shown in Fig. 1.
Cyclin-dependent kinases: an entbarrassntent of riches?
Authors: Pines Jonathon , Hunter Tony 
Published: November 2023

Abstract
One of the most exciting advances in our understanding of the cell cycle has been the identification in multicellular organisms of a family of protein kinases that are involved in the regulation of different checkpoints. These protein kinases are related both in their primary structure, and because they bind and are activated by the cyclin family of proteins. The latter property has led to this family of protein kinases being called the ‘cyclin-dependent kinases’, or cdks. The cyclins are a family of structurally related proteins that typically vary in amount during the cell cycle in a programmed manner (the cyclins are reviewed in Chapter 5). The first two cyclins to be isolated, cyclins A and B, were called mitotic cyclins, because they were specifically destroyed in mitosis, implicating them in the regulation of the G2/M transition. Subsequently, another class of cyclins was discovered in yeast that have roles in the Gl/S transition, and are called Gl cyclins. More than ten different cyclins have been identified in humans and in budding yeast, and the different cyclin/cdk complexes are involved in regulating several different check –points throughout the cell cycle. Furthermore, it now appears that some cyclin –cdk complexes regulate processes quite separate from the cell cycle. The paradigm for the cyclin-cdk interaction is that between cdc2 and cyclin B (reviewed in Chapter 5). Therefore, in this article we will only outline the salient points.
The packaging of pre-Mrna
Authors: Mcafee James g. , huang mae , soltaninassab syrus , Rech janee. , iyengar, sunita , Lestourgeon wallace m. 
Published: November 2023

Abstract
In eukaryotes, pre-mRNA is bound by protein either simultaneously with the initiation of transcription or very soon thereafter (1-3), and it is not possible to isolate pre-mRNA molecules free from a unique set of abundant nuclear proteins unless protein denaturants are used in the purification scheme. The packaging of DNA into nucleosomes, and their association in the 30 nm chromatin fibre, greatly foreshortens the DNA substrate and provides some protection of the genome from nuclease activity (4). However, the major pre-mRNA binding proteins possess well- characterized, helix-unwinding activities (5-8) and, in comparison with nucleosome packaging, provide less protection from nuclease activity (9,10). It seems then, that the pre-mRNA packaging mechanism functions primarily to maintain the transcript in a single-stranded state. This generally fits with the short-lived nature of pre- mRNA, and with the fact that nascent transcripts must be accessed by factors involved in intron removal, polyadenylation, and transport. Unlike rRNA, tRNA, and snRNA, pre-mRNAs are usually transcribed from single-copy genes and the pool of total nuclear pre-mRNA reflects great length heterogeneity (hence the term, heterogeneous nuclear RNA or hnRNA).
Spliceosome assembly
Authors: Reed Robin , Palandjian Leon 
Published: November 2023

Abstract
Pre-mRNA splicing takes place within a large, highly dynamic complex designated the spliceosome. Spliceosomes are 50-60S, have a calculated molecular weight on the order of 3-5 × 106 kDa, and are estimated to be 40-60 nm in diameter (see 1, 2 for reviews). Among the best characterized of the spliceosomal components are the small nuclear RNAs (snRNAs) Ul, U2, U4, U5, and U6, which are thought to play central roles not only in spliceosome assembly, but also in the two catalytic steps of the splicing reaction (reviewed in 1, 3, 4; see Chapter 5). Less is known about the protein components of the splicing machinery. Genetic studies in yeast have identified over 50 protein-coding genes essential for splicing (reviewed in 2; see Chapter 7), and an equal number of proteins have been detected in highly purified mammalian spliceosomes (5, 6). In addition to obtaining direct evidence for RNA- mediated catalysis, major challenges of the splicing field include cloning, characterizing, and determining the functions of the protein components of the spliceosome and achieving a detailed understanding of spliceosome assembly.
snRNP structure and function
Authors: Will Cindy l. , lÜhrmann reinhard 
Published: November 2023

Abstract
In contrast to self-splicing RNA molecules which contain extensive, highly con¬ served structural elements sufficient for intron excision and exon ligation, nuclear pre-mRNA molecules possess short, conserved sequences which are limited to the 5’ and 3’ splice site and branchpoint regions (see Chapters 1, 4, and 8). While these sequences are necessary for the conversion of the pre-mRNA to mRNA, they are not sufficient. Nuclear pre-mRNA splicing is thus dependent upon the activity of a number of trans-acting splicing factors, which in addition to protein factors, include the four small nuclear ribonucleoprotein particles (snRNPs) Ul, U2, US, and U4/U6. These evolutionarily conserved RNA-protein complexes interact stepwise with nuclear pre-mRNA molecules to form the large multicomponent RNP complex (the spliceosome) in which splicing occurs (see Chapter 4). Ul, U2, US, and U4/U6 snRNPs play a central role in the recognition and alignment of splice sites. In addition, snRNPs may directly contribute to the catalytic activity of spliceosomes: snRNA constituents have been proposed to form the catalytic centre responsible for excision-ligation events.
Mammalian pre-mRNA splicing factors
Authors: Caceres Javier f. , Krainer adrian r. 
Published: November 2023

Abstract
Since the discovery of pre-mRNA splicing in 1977, considerable progress has been made in dissecting the czs-elements and frzms-acting factors involved in carrying out this complex reaction. Multiple protein and nucleoprotein components are necessary to catalyse pre-mRNA splicing, which occurs within a particle termed the spliceosome. The components of the splicing machinery, and/or factors involved in the assembly of the spliceosome, include the small nuclear ribonucleoprotein particles (snRNPs) Ul, U2, U4/U6, and U5; the polypeptides that associate with hnRNA to form hnRNP particles (hnRNP proteins); and a large number of non- snRNP essential splicing factors (for earlier reviews see 1-6).
Alternative pre-mRNA splicing
Authors: wang Yung-chih , selvakumar, meenakshi , Helfman david 
Published: November 2023

Abstract
The formation of a mature mRNA from a primary transcript generally requires excision of intervening sequences (introns) with the subsequent joining together (splicing) of exons. The general splicing reaction of metazoan intrans has been well characterized and involves a two-step process, which occurs in a large ribonuclear protein complex, termed the spliceosome (see Chapter 4; reviewed in 1, 2). In the first step, a 5’ exon is cleaved and two intermediates are formed: a 5’ exon with a 3’ hydroxyl end, and an RNA molecule containing the downstream intron sequence and 3’ exon, in which the 5’ terminal guanosine of the intron is covalently linked via a 2’-5’-phosphodiester bond to a residue termed the branchpoint, usually an adenosine, located 18-40 nucleotides upstream of the 3’ splice site. In the second step, cleavage at the 3’ splice site results in the release of a lariat intron and con-comitantly the two exons are ligated together.
Analysis of Gene Expression by Reverse Transcriptase-Polymerase Chain Reaction
Authors: Koopman Peter 
Published: November 2023

Abstract
A molecular genetic approach to development often involves isolating genes controlling cellular developmental processes, analysing their structure and regulation, and gaining clues about their function by examining their expression. Two constraints commonly encountered in studying gene expression in embryos are scarcity of tissue and low levels of expression. When the gene of interest is expressed in small groups of cells, or when transcripts are present in vanishingly low amounts, it is difficult to perform Northern blotting, RNAase protection assays, or in situ hybridization. In these cases the exquisite sensitivity of the polymerase chain reaction (PCR) can provide a solution. PCR has been used to detect transcripts present at down to one copy per thousand cells (1), and in samples as small as a mouse blastocyst (2-4), or even a single cell (5, 6).
Oligonucleotide-directed mutagenesis with single-stranded cloning vectors
Authors: Kolmar Harald , Fritz Hans-Joachim 
Published: November 2023

Abstract
Directed mutagenesis with synthetic oligonucleotides was proposed in 1971 by Hutchison and Edgell (1) and first developed to a practical procedure by Zoller and Smith (2, 3). According to the original protocol, the gene of interest is first cloned in a vector derived from phage M13 (single-stranded circular DNA genome). An oligonucleotide is synthesized that is complementary to the target site of mutagenesis with the exception of one or more non complementary nucleotide central residues. The oligonucleotide is annealed to the single-stranded, recombinant vector DNA, and covalently closed double-stranded heteroduplex DNA (hd DNA) containing a single site of non-homology is synthesized in an enzyme catalysed reaction. This hd DNA is transfected into E. coli where it replicates in vivo, to segregate DNA clones carrying the predetermined mutation carried on the synthetic oligonucleotide. Some of the first spectacular successes of protein engineering were achieved with this method (see for example ref. 4). It was soon realized, however, that the procedure is bound to produce only low yields of the desired mutants because cellular processes of DNA mismatch repair are directed against the DNA strand synthesized in vitro (5). Marker yield is indeed an important aspect, since it drastically influences the work needed to identify a clone with the correct structure, which-in the absence of a phenotype-has to be accomplished solely by biochemical means such as nucleotide sequence analysis (5).
Expression of cloned genes in yeast
Authors: Romanos Michael A , Scorer Carol A , Clare Andjeffrey J 
Published: November 2023

Abstract
Saccharomyces cerevisiae has long been a model organism for biochemical and genetic studies, and the expression of cloned genes in yeast was examined as soon as transformation procedures became available. The particular advantages it offers over other expression systems are mainly due to the fact that it is both a micro-organism and a eukaryote. Its genetics are more advanced than any other eukaryote, so that it can be genetically manipulated almost as easily and rapidly as Escherichia coli. As a eukaryote, it is more likely to provide a suitable environment for the folding of foreign eukaryotic proteins than E. coli, and can be more readily used for secretion of proteins. Yeast can be grown rapidly on simple media, and to high cell density. Additionally, as a food organism, yeast is regarded as highly acceptable for the production of pharmaceutical proteins.
Expression using a defective herpes simplex virus (HSV-1) vector system
Authors: Lim Filip , Starr Philip , Song Song , Hartley Dean , Lang Phung , Wang yaming , Geller Alfred I 
Published: November 2023

Abstract
Genetic intervention in the brain through the use of vector systems capable of transferring recombinant genes into mammalian neurones is becoming an increasingly useful strategy for investigating the molecular basis of various brain functions ranging from regulating homeostasis to information processing and higher cognitive functions (1, 2). Genes have been isolated and characterized which encode molecules such as signal transduction enzymes, transcription factors, and components of the neurotransmitter release machinery that may play critical roles in regulating neuronal physiology. Relatively little is known about the function of these molecules in mediating alterations in neuronal physiology and the resulting changes in complex behaviours. Genetic intervention in the brain has the theoretical potential to reveal precise information about specific brain functions through the use of mutations in neuronal genes, and then subsequent expression of the altered gene products in their normal cellular milieu.
Cosmid clones and their application to genome studies
Authors: Ivens Alasdair c. , Little Peter F R 
Published: November 2023

Abstract
A 1000-fold range of DNA sizes may be cloned in the current range of cloning vectors: the ideal vector for genome mapping studies would be one that is easy to use while containing as much DNA as possible. Cosmids now occupy the middle ground: they have a significant capacity, thus reducing the number of steps required to clone an entire gene or region, combined with very simple methods for isolating inserted DNA in pure form. As a consequence, positional cloning strategies frequently involve the use of cosmids as a final cloning vector for reducing a yeast artificial chromosome (YAC) clone to manageably sized DNA fragments (1). Cosmids are units that are very likely to contain an entire gene, are easily mapped with respect to restriction sites, and are amenable to the application of a number of other functional assays, e.g. exon trapping (2, 3), genomic sequencing (4-6), and fingerprinting to generate contig maps (7, 8). As a result, the detailed information that can be obtained from a cosmid clone makes it the ideal medium for genome analysis. Indeed, several genome mapping studies (e.g, Caenorhabditis elegans (9), Escherichia coli (10)), have relied on physical DNA maps built around cosmid clones.
Isolation and characterization of photosynthetic reaction centres from eukaryotic organisms
Authors: Evans Michael c.w. , Hallahan Beverly j. , Hanley Jonathan A , Heathcote Peter , Gumel Nicola J , Saul Purton 
Published: November 2023

Abstract
Photosynthetic reaction centres are the site of solar energy conversion in biological systems. The basic organization of photosynthetic systems is the same in all chlorophyll-containing organisms. Light is absorbed by a large array of chlorophyll molecules and the energy transferred by random processes through the array to a reaction centre chlorophyll, which undergoes photochemical oxidation transferring an electron to an acceptor molecule. The chlorophyll array is highly organized with the pigments bound to a range of proteins which modulate the chlorophyll energy levels and provide an organization which facilitates energy transfer to the reaction centre. The light harvesting protein complexes surround the reaction centre forming a photosynthetic unit. The photosynthetic units are in tum bound in a specific membrane system, the thylakoid. In eukaryotic organisms the thylakoids are in tum limited to a specialized organelle, the chloroplast. The high chlorophyll to reaction centre concentration makes optical analysis of the reaction centre extremely difficult and also limits the concentration of reaction centres which can be obtained for spectroscopic analysis. The presence of two types of reaction centre in oxygen evolving organisms further complicates analysis. The successful development of models of reaction centre structure and mechanism has depended on the development of techniques to isolate and characterize the reaction centres.
Haemolytic assays for whole complement activity and individual components
Authors: Whaley Keith , North Jonathan 
Published: November 2023

Abstract
The ability of the intact classical or alternative pathways of complement to lyse erythrocytes is used in functional assays of entire pathways or of single components. Functional assays have the advantage over immunochemical assays of individual components, as the latter, depending on the choice of detecting antibody, can measure non-functional proteins and breakdown products generated by complement activation in addition to the native proteins. Reproducible haemolytic assays depend on performing the tests in conditions similar to those in vivo in respect of isotonicity and pH, as deviations from this can lead to spontaneous lysis of the target cell.
Purification of complement components
Authors: Daha Mohamed R 
Published: November 2023

Abstract
Purified complement components are required for many purposes, including assessment of their biological activities, use as standards and reagents in assays, and raising of antibodies. In particular, the role of complement in vivo requires further extensive research in the near future. It is thought that transgenic and knockout models of complement may provide valuable additional information, and in these studies the substitution of missing complement components with purified reagents may prove to be essential.
Biosynthesis of complement components
Authors: Circolo Antonella , Nutter Thomas B , Strunk Robert C 
Published: November 2023

Abstract
Complement protein biosynthesis has been studied since the 1890s when Metchnikov suggested that leucocytes were a source of complement (1). Ehrlich and Morgenroth (2) were the first to suggest that the liver was a site of complement production when they demonstrated that complement, but not antibody, was depressed in sera of animals poisoned with phosphorus. In the late 1960s, Alper and co-workers (3) confirmed that the liver was a major site of serum complement synthesis by studying a patient following liver transplantation; the shift of C3 from recipient to donor type indicated that the liver synthesized at least 95% of the C3 found in serum.
Arabidopsis mutant characterization; microscopy, mapping, and gene expression analysis
Authors: Kalantidis Kriton , Greg Briarty L , Wilson Zoe A 
Published: November 2023

Abstract
Mutants provide a valuable resource for the study of developmental biology, since the effects of genes can be identified and their role in development assessed in homozygous and heterozygous states. One problem with gene identification by mutagenesis can be the cloning of the specific gene (see Chapters 7 and 8). This chapter will cover a range of approaches which can be used to characterize mutants of Arabidopsis thaliana. These include the initial mapping of the mutation and determination of allelism between other mutations with similar phenotypes, both by classical linkage analysis and by molecular approaches. Different approaches are presented to allow the detailed analysis of phenotype by microscopy. The final part of the chapter gives protocols to permit the analysis of gene expression by Northern and in situ hybridization.
Tissue culture, transformation, and transient gene expression in Arabidopsis
Authors: Lindsey Keith , Wei Wenbin 
Published: November 2023

Abstract
Tissue culture and transformation technologies have revolutionized studies on plant biology. They allow investigations, in a way previously only dreamed of, of the relationships between, plant morphology on the one hand and gene expression, metabolism, and cell differentiation on the other. Disorganized growth, such as occurs during callus formation, can be experimentally induced and the developmental, biochemical, and genetic consequences studied (1). Furthermore, and of fundamental significance, disorganized tissues can be induced to reorganize and to regenerate intact organs and whole plants. This observation has been crucial in the development of the concept of the totipotency of plant cells, and has been equally fundamental to the establishment of reproducible techniques for the genetic engineering of plants. Thus, it is possible to introduce foreign genes into cultured plant cells, protoplasts, or tissues, and regenerate from them stably transformed individuals that will transmit the introduced transgenes to their progeny. It is beyond the scope of this chapter to discuss the commercial or agronomic implications of plant genetic engineering, but applications that are of a more basic scientific nature, and to which Arabidopsis thaliana lends itself ideally as an experimental organism, can briefly be considered here.
Cell cycle analysis by flow cytometry
Authors: Crissman Harry A 
Published: November 2023

Abstract
Mechanisms that control and regulate the cell cycle are currently being intensively studied in numerous laboratories. Central to these studies is the basic understanding of those regulatory mechanisms that appear to be lost or dysfunctional when cells become transformed from the normal to the malignant state. This information can lead to a better understanding of the nature of cancer and potentially result in the design of more effective chemotherapeutic agents or even to the intervention of those processes that lead to the development of cancer. Studies in these areas will necessarily rely on the constant assessment of cell cycle distribution and cycle progression.
Analysis of flow cytometric DNA histograms
Authors: Rabinovitch Peters. 
Published: November 2023

Abstract
Analysis of the DNA content of cells and their distribution within G1, S and G2/M phases of the cell cycle is one of the earliest applications of flow cytometry. Because of its simplicity, DNA content analysis remains a commonly used and powerful technique. Chapters 2 and 8 within this volume describe additional variations and enhancements of the original flow cytometric techniques; however, mathematical analysis of single parameter DNA content histograms remains a common element in many of these methods. Recent interest in the clinical application of DNA ploidy and cell cycle measurements from DNA histograms has stimulated improvements in the methods and models used to derive these measurements, and guidelines for the implementation of DNA cytometry have been established to help ensure greater accuracy in implementation of these techniques. This chapter describes a protocol consistent with these guidelines for DNA content histogram analysis and discusses some of the principles in methods of analysis.
Morphological and biochemical criteria of apoptosis
Authors: Ramachandra Sumant , Studzinski George P 
Published: November 2023

Abstract
When cells receive mixed signals for growth they usually die. For instance, when the developmental programme requires cell division but external growth signals are lacking, or when a growth-related gene such as c-myc is highly expressed but the cellular environment has insufficient nutrient content, or a xenobiotic is present, the cell dies by a process termed apoptosis. Although there are differences in the phenomena observed during this sequence of events, depending on the cell type, and agent or circumstance which initiates the cell’s demise, there are morphological and biochemical similarities which suggest that these are variants of the same biological process designed to control the size of cell populations.