bluuebunny/crossref_metadata_2025_split huggingface dataset:
Polymerase chain reaction‐based assays for the characterisation of plant genetic resources
Authors: Powell Wayne , Orozco‐Castillo Carlos , Chalmers Kenneth J. , Provan Jim , Waugh Robbie 
Published: May 2005

AbstractPlant genetic resources are an important component of biodiversity and provide the basic genetic variability that allow new and improved cultivars to be developed. Numerous germplasm collections have been established and it is important to establish that such collections are representative and accessible to breeders and biotechnologists. Molecular markers provide the best estimate of genetic diversity since they are independent of the confounding effects of environmental factors. Assays based on the polymerase chain reaction (PCR) are considered to meet both the technical and genetical requirements for the characterisation of plant and animal genetic resources. Two main approaches are described, based on anonymous and defined primers. The use of both randomly amplified polymorphic DNA (RAPD) and microsatellites or simple sequence repeats (SSR) for the characterisation of perennial tree species, and distribution of variability within gene pools is reported. The detection of interspecific gene introgression between coffee species with RAPD markers is described together with the use of microsatellites to genotype potato. The use of PCR‐based assays will facilitate the evaluation and utilisation of plant genetic resources.
DNA profiling of banana and plantain cultivars using random amplified polymorphic DNA (RAPD) and restriction fragment length polymorphism (RFLP) markers
Authors: Bhat K. Venkataramana , Jarret Robert L. , Rana Rai Singh 
Published: May 2005

AbstractPolymerase chain reaction (PCR) amplification of genomic DNA from 57 Musa cultivars with 60 random 10‐mer primers generated 605 polymorphic amplification products which were useful in unambiguous cultivar identifications. Unweighted pair‐group method analysis of this data grouped the cultivars into specific clusters depending on their genomic similarities. The diploid ancestral species of cultivated banana and plantains, namely Musa acuminata ssp malaccensis, an A genome donor and M. balbisiana, a B genome donor, were farthest apart from each other in the phenogram. The edible fruit yielding cultivars with the genomic constitutions AA, AAA, AB, AAB, ABB and ABBB grouped in different clusters according to overall genetic homologies. The restriction fragment length polymorphisms (RFLPs) prevalent among the cultivars were studied by hybridizations of 19 random genomic clones to blots of HindIII, EcoRI and MspI digests. Cluster analysis of these data on 107 polymorphic alleles resulted in a phenogram comparable to the one obtained with random amplified polymorphic DNA (RAPD) analysis. Two multilocus probes useful in distinguishing all the 57 cultivars analyzed were also identified. The A and B types of cytoplasms in the cultivars were further distinguished by hybridization of heterologous chloroplast DNA probes. Results showed that use of different kinds of molecular markers in gene banks is essential for characterization and classification of germplasm collections.
Protein variability in male and female Wistar rat liver proteins
Authors: Steiner Sandra , Wahl Daniel , del Carmen Varela Maria , Aicher Lothar , Prieto Pilar 
Published: May 2005

AbstractIn the emerging field of mechanistic toxicology, growing attention is being paid to the interpretation of sex‐related toxic responses. The Wistar rat outbred stock is a frequently used rodent for toxicity testing. Outbred strains, which display a relatively high degree of genetic variability between the individual animals are often used in risk assessment, as they are considered to best approximate the variability present in a human population. In this study two‐dimensional (2‐D) protein gel electrophoresis was applied to investigate the liver protein patterns of male and female Wistar rats and to search for (i) sex‐related and (ii) interindividual qualitative and quantitative differences in protein expression. Among the sexes, six proteins were detected that were shown to be exclusively present in male rats and one that was present only in females. A male‐specific protein was tentatively assigned to α2u globulin. Seven protein spots showed statistically significant abundance changes (p < 0.001) between males and females, one was tentatively assigned to heme oxygenase 1 and another to the 23 kDa morphin‐binding protein. Four sets of protein spots were detected that showed positional shifts in the individual patterns and are likely to represent polymorphic proteins inherent in the Wistar rat. These results form a valuable basis for future investigations of drug‐induced changes in the male and female Wistar rat liver pattern.
Sample self‐stacking and sample stacking in zone electrophoresis with major sample components of like charge: General model and scheme of possible modes
Authors: Gebauer Petr , Thormann Wolfgang , Boček Petr 
Published: May 2005

AbstractA theoretical study is presented of zone electrophoretic behavior of samples that contain one or more minor analytes and at least one major ionogenic component of like charge. Based on a simple model comprising weak univalent anionic electrolytes, conditions are derived under which analytes are temporarily focused isotachophoretically into very narrow zones by a sample self‐stacking effect provided by the major sample components. Requirements for minimal/maximal mobility and a background coion concentration dependent minimal concentration of a major sample component (stacker) are presented. For systems in which sample self‐stacking does not apply, an expression for the concentrating factor is derived that involves the effects of both nonselective (classical) and selective sample stacking, the latter being a consequence of electrophoretic separation of the minor analyte from the major component. The theory derived is discussed with selected model examples by using both numerical calculation and computer simulation.
Demethylation of a repetitive DNA sequence in human cancers
Authors: Miwa Wataru , Yashima Kazuo , Sekine Teruaki , Sekiya Takao 
Published: May 2005

AbstractTo detect DNA alterations in unknown regions in human cancers, we have performed restriction landmark genomic scanning (RLGS) analysis of DNA isolated from cancer and normal cells. One spot with a highly intensified signal was detected in DNA from all six malignant melanoma cell lines, two of five colon cancer cell lines and one of six pancreatic cancer cell lines analyzed. In DNA from normal cells, two placentas and seven cultured lymphocytes, the signal of this spot was not intense. The DNA fragment corresponding to the spot was cloned. By nucleotide sequence analysis, the DNA fragment was revealed to be a part of a repeating unit of a 13 kbp nucleotide sequence of which 200 copies were located in chromosome 8q21. Southern blotting analysis using the cloned fragment as a probe demonstrated that the intensified signal for the DNA fragment observed in cancer cells was due to demethylation in the recognition sequence of the NotI restriction enzyme. The results suggest that marked demethylation in the repeating units might be associated with the genesis or progression of some types of cancers.
Application of fluorescence in situ hybridization in genome analysis of the mouse
Authors: Matsuda Yoichi , Chapman Verne M. 
Published: May 2005

AbstractFluorescence in situ hybridization (FISH) is an effective technique for localizing cloned DNA probes directly onto metaphase chromosomes. Human genome mapping using FISH has been significantly enhanced by the development of new techniques, especially high‐resolution gene mapping with direct R‐banding FISH and physical gene ordering with multi‐color FISH. By contrast, FISH techniques have not been put to practical‐use for the analysis of the mouse genome compared with the human. We have developed and modified FISH techniques for use in mouse genome analysis. In this article we summarize and review our recent results with FISH analyses in the following studies: (i) high‐resolution gene mapping with the direct R‐banding FISH, (ii) analysis of chromosomal rearrangement with multi‐color FISH, (iii) establishment of centromere mapping with the major satellite DNA probe (iv) analysis of chromatin structure in meiotic cells, and (v) application of FISH in cytogenetic studies of genetic variation in the mouse, showing that these applications of FISH are very useful for mouse genome analysis.
Genomic mismatch scanning: Current progress and potential applications
Authors: Nelson Stanley F. 
Published: May 2005

AbstractGenomic mismatch scanning (GMS) is a new method of genetic mapping which attempts to purify and map the regions of identity between two complex genomes in a single test. Identical DNA fragments from two genomic sources are enriched in two steps: (i) after reannealing of the two genomes, heterohybrids are purified by using a combination of a restriction methylase and methylation‐sensitive endonucleases, (ii) heterohybrids that contain mismatches are nicked in vitro by the E. coli MutHLS mismatch repair system and are eliminated subsequently from the pool, leaving only mismatch‐free heterohybrids. The genomic origin of this selected pool of DNA fragments is then mapped in a single hybridization step onto metaphase chromosomes or ordered DNA arrays. The principal advantages of GMS are (i) it approaches the theoretical limit of mapping power and resolution offered by an arbitrarily dense set of completely informative polymorphic markers and (ii) it results in a great increase in the effective number of informative markers without a corresponding increase in the number of individual tests. Thus, it should provide an efficient method for affected‐relative‐pair linkage mapping and for linkage disequilibrium mapping. In addition, a variation of GMS may allow rapid genomic scanning for regions of homozygosity‐by‐descent or somatic loss‐of‐heterozygosity. The feasibility of GMS has been validated in the 15 mb genome of Saccharomyces cerevisiae. This article discusses the principles of GMS, the application to more complex genomes, and the possible uses of GMS.
Rapid and effective Western blotting of histones from acid‐urea‐Triton and sodium docecyl sulfate polyacrylamide gels: Two different approaches depending on the subsequent qualitative or quantitative analysis
Authors: Thiriet Christophe , Albert Philippe 
Published: May 2005

AbstractAn improved method for the electrophoretic transfer of histones from sodium dodecyl sulfate (SDS) and acetic acid‐urea‐Triton X‐100 (AUT) polyacrylamide gels onto nitrocellulose membranes is described. In the case of SDS‐gels, it was not essential to equilibrate them before transfer while for the AUT‐gels, an equilibration step is essential to prevent the interference of Triton X‐100 with the binding of histones to nitrocellulose. Transfer efficiency was different for different histone classes. Two procedures were developed: (i) one suitable for qualitative studies, and (ii) another for quantitative transfer.
Separation and detection of amino acids and their enantiomers by capillary electrophoresis: A review
Authors: Issaq Haleem J. , Chan King C. 
Published: May 2005

AbstractSince its introduction as an analytical technique capillary electrophoresis has been used for the separation of amino acids and their enantiomers; over 150 studies have been published to date. This review deals with their separation and detection. Amino acids have been resolved using both capillary zone electrophoresis and micellar electrokinetic chromatography. Pre‐column derivatization schemes which are employed for the sensitive detection of amino acids are discussed. Criteria for the selection of the pre‐ or post‐column derivatizing agent, chromophore or fluorophore, are presented. Detection systems, direct and indirect, that have been used are given with emphasis on fluorogenic reagents and laser induced fluorescence detection. Also, procedures for the separation of amino acid enantiomers are discussed and illustrated.
Enantiomeric separation of amino acids using micellar electrokinetic chromatography after pre‐column derivatization with the chiral reagent 1‐(9‐fluorenyl)‐ethyl chloroformate
Authors: Chan King C. , Muschik Gary M. , Issaq Haleem J. 
Published: May 2005

AbstractDirect enantiomeric separations of some racemic amino acids derivatized with 9‐fluorenylmethyl chloroformate were obtained using cyclodextrin‐modified micellar electrokinetic chromatography (CD/MEKC) with a buffer made up of 5 mM sodium borate (pH 9.2), 150 mM sodium dodecyl sulfate (SDS) and 40 mM γ‐CD. Alternatively, enantiomeric separations were also achieved indirectly using MEKC after pre‐column derivatization with (+)‐1‐(9‐fluorenyl) ethyl chloroformate (FLEC). Using either a 10 mM sodium phosphate (pH 6.8) or a 5 mM sodium borate buffer (pH 9.2), each of which contained 25 mM SDS and 10–15% of acetonitrile, FLEC‐derivatized serine, alanine, valine, methionine, leucine, phenylalanine, tryptophan, and their diastereomeric pairs were all separated: the L‐isomers migrated faster than the corresponding D‐isomers. However, when (−)‐FLEC was used for derivatization, the D‐isomers migrated faster than the corresponding L‐isomers. Also, the diastereomers of aspartic acid, glutamic acid, and proline were resolved using a 10 mM sodium citrate buffer (pH 4.4). Using KrF (248 nm) laser‐induced fluorescence, the detection limit of (+)‐FLEC derivatized DL‐amino acids was obtained at the nM level, which was about 100 × more sensitive than UV absorption at 200 nm. Analyte concentrations as low as 3 × 10−8 M (DL‐Val) could be derivatized with (+)‐FLEC.
Application of capillary electrophoresis to amino acid sequencing of peptide
Authors: Kim Nam Jung , Kim Jung Ho , Lee Kong‐Joo 
Published: May 2005

AbstractSeparations of twenty phenylthiohydantoin (PTH) amino acids, and amino acid derivatives, resulting from the Edman degradation of peptides and proteins, were optimized for peptide sequencing by capillary electrophoresis. Manual sequencing of angiotensin II was performed by Edman degradation and capillary electrophoresis of the PTH amino acid obtained after each cycle. The results were compared with those of an automated conventional protein sequencer. Interfacing capillary electrophoresis with Edman degradation provides an additional option for protein sequencing.
Fast, high‐resolution (capillary) electrophoresis in buffers designed for high field strengths
Authors: Hjertén Stellan , Valtcheva Ludmila , Elenbring Karin , Liao Jia‐Li 
Published: May 2005

AbstractCapillary electrophoresis in conventional buffers and in 50 μ capillaries permits field strengths as high as 300–500 V/cm with acceptably low thermal zone deformation. However, still higher field strengths (up to at least 2000 V/cm) can be applied without a decrease in resolution if the experiments are performed in the buffers described in this paper. Characteristic of these buffers is their low electrical conductivity and yet satisfactory buffering capacity accomplished either (i) by selecting buffer constituents of relatively high molecular weight and small net charge or (ii) by fractionation of carrier ampholytes (originally introduced for isoelectric focusing experiments) into a series of narrow pH range fractions and using these fractions as buffers, or (iii) by selecting an ampholyte with two acidic groups and one basic group (or one acidic group and two basic groups) and with a pI value close to two of its pK values. In such buffers, aromatic carboxylic acids and proteins used as model substances could be analyzed rapidly. For instance, albumin and transferrin were separated at 30000 V (1.99 μA) in 15 cm long fused silica capillaries (50 μ ID) within 40 s and the carboxylic acids within 25 s. The resolution was similar to that obtained at standard voltage (5000 V; 0.33 μA), but the analysis time was reduced sixfold. Although not verified experimentally we also suggest the use of relatively high‐molecular‐weight polyoxyethylene derivatized with one acidic group (for instance, boric acid) and one basic group (an amine), both having the same pK value, which should afford both a very high buffering capacity and very low electrical conductivity (at low buffer concentrations).
Effect of buffer composition on the migration order and separation of histone H1 subtypes
Authors: Lindner Herbert , Helliger Wilfried , Sarg Bettina , Meraner Christoph 
Published: May 2005

AbstractThe effects of different buffer concentrations and compositions on the elution order and separation of H1 histone subtypes and their phosphorylated modifications isolated from several species was studied using high‐performance capillary electrophoresis (CE). Various cations and anions were tested in an untreated silica capillary and low pH buffers, in the presence of the dynamic coating agent hydroxypropylmethyl cellulose. It was found that the cations and anions of buffers have a remarkable influence on both the efficiency and the selectivity of protein separations. A triethylammonium methanephosphonate system proved efficacious for the separation of rat histone subtype H1c from H1e and a perchlorate/triethylammonium phosphate system for the analysis of chicken and mouse linker histones. CE provides an attractive alternative to high‐performance liquid chromatography and conventional gel electrophoresis.
Ribbon channel plate rotating drum DNA sequencing device
Authors: Douthart Richard J. , Welt Marc , Walling Leslie 
Published: May 2005

AbstractA new design DNA sequencing electrophoresis device is described. The device, called the ribbon channeled plate rotating drum (rprd), consists of two major components, the plate assembly and the drum assembly. The plate assembly contains a machined or etched plate of individual micro‐channels called the ribbon channeled plate. The ribbon channeled plate and other components of the plate assembly combine the advantages of thin gels and capillary arrays in a single unit with few of the disadvantages. The other major component of rprd is the drum assembly, which facilitates direct blotting onto deposition membranes affixed to a large plastic drum. The drum with attached membrane and deposited electrophoretically resolved ladders is easily moved to special units facilitating downstream processing and detection. The drum unit, although versatile, is specifically designed to be used with multiplex sequencing.
The use of a nonradioactively labelled probe system in an electrophoretic ribotyping method for the differentiation of strains of coagulase‐negative staphylococci
Authors: Hawcroft David M. , Geary Colin 
Published: May 2005

AbstractBacteria of the genus Staphylococcus are notoriously difficult to classify and identify; many routine hospital laboratories use only simple, older techniques to segregate them into broad groups. Recent studies have demonstrated that subspecific strains vary in their clinical significance and are of increasing importance in hospital‐derived infections; this has led to a need for better methods of classification and identification. Restriction fragment length polymorphism analysis of genes for ribosomal RNA (‘ribotyping’) is potentially very useful since these genes occur in multiple copies in the genome, and are relatively stable. The restriction fragment patterns are sufficiently different to be usable in classification and identification. This paper considers an electrophoretic method for the separation of these restriction gragments which has the advantages of using a nonradioactive probe and a simple colour signal generating system for its evaluation. The technical principles are universal and would allow the procedure to be developed for other groups of microorganisms.
Glycoproteins in human parotid saliva assessed by lectin probes after resolution by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis
Authors: Carpenter Guy H. , Proctor Gordon B. , Pankhurst Caroline L. , Linden Roger W. , Shori Deepak K. , Zhang Xiao S. 
Published: May 2005

AbstractHuman parotid salivary glycoproteins separated by gradient sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and electroblotted onto nitrocellulose have been investigated using a battery of biotinylated lectin probes of characterized sugar specificity. Lectin binding, detected on blots using avidin‐biotin complex (ABC) and a chemiluminescence generating substrate, was recorded on photographic film and compared with the original fluorescein isothiocyanate (FITC) stained blots or with Coomassie Brilliant Blue R‐250‐stained gels run in parallel. A number of glycoprotein bands which were undetected by protein stains or the periodic acid Schiff reaction were revealed by lectins. Binding by lectins from Concanavalia ensiformis, Lens culinaris, Limax flavus, Phaseolus vulgaris, Ricinus communis, Triticum vulgaris, Lotus tetragonobulus and Ulex europaeus indicated that sialylated and fucosylated triantennary and bisected, N‐linked complex sugar chains were present on many glycoproteins in addition to the major glycosylated proline‐rich glycoprotein (GI). Binding with lectins from Arachis hypogaea and Dolichos biflorus indicated that the O‐linked sugar chains were confined to the α‐heavy chain of Ig A. Comparison of lectin binding in samples from five healthy individuals revealed differences in a number of flycoproteins in addition to the previously characterized GI and CON 1/CON 2 polymorphisms and demonstrated that the H blood group antigen was expressed mainly on GI in parotid saliva. This study will be used as a basis upon which to study salivary glycoproteins in diseases affecting parotid glands.
Analysis of heparin‐like pharmaceuticals by capillary zone electrophoresis and isotachophoresis
Authors: Malá Zdeňka , Křváková Ludmila , Boček Petr 
Published: May 2005

AbstractSynthetic sulfated bis‐lactobionic acid amides are important heparin‐like pharmaceuticals. The synthesis of these compounds yields molecules differing in the number of sulfate groups, and during the isolation procedure, the required species may partially decompose or take in some impurities. This article shows that capillary zone electrophoresis may serve well as an expedient method for the analysis of the above‐mentioned pharmaceuticals. Complex‐forming equilibria between the analytes and bivalent cations present in the background electrolyte bring selectivity necessary for the separation, and the detection at various wavelengths serves as an aid in the characterization of admixtures, decomposition products, and impurities. Capillary isotachophoresis may also be used for the analysis of these species, bringing about the potential of micropreparation of individual compounds and opening the chance for continuous free‐flow electrophoresis.
Dextrin sulfopropyl ether: A novel anionic chiral buffer additive for enantiomer separation by electrokinetic chromatography
Authors: Jung Martin , Börnsen K. Olaf , Francotte Eric 
Published: May 2005

AbstractDextrin 10 sulfopropyl ether (DSPE) was prepared and characterized matrix‐assisted laser‐desorption ionization‐mass spectrometry (MALDI‐M) using a recently developed matrix. The anionic compound was used as a novel chiral buffer additive for enantiomer separation by capillary electrophoresis, thereby changing into the elektrokinetic chromatography (EKC) mode. DSPE was systematically evaluated as a chiral selector and was compared to the respective nonderivatized maltodextrin. DSPE showed an increased separation power for cationic racemic solutes. Although not quite as versatile and powerful as cyclodextrins, the inexpensive dextrin 10 and its derivative DSPE showed remarkable enantioselectivity in some cases.