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C J Bostock

Publications and source records attributed to C J Bostock.

At least 37 records · Page 2Linked to original sources
PubMed1990

Detection of phocine distemper virus using the polymerase chain reaction.

During the fatal seal epizootics in the North and Baltic Seas in summer 1988 a virus was isolated which was shown to be the causal agent. It was subsequently classified as morbillivirus by neutralization assays, reaction with monoclonal antibodies and nucleic acid hybridization studies. The virus (tentatively called Phocine Distemper Virus, PDV) is difficult to grow in culture making rapid diagnosis difficult. We have used the Polymerase Chain Reaction (PCR) as an alternative and fast method to detect the presence of virus-specific nucleic acid and we describe here the amplification of cell culture derived PDV RNA in a "one-tube" reaction using heterologous (Rinderpest Virus cDNA derived) F gene primers. The resulting 370 bp DNA fragment was shown to be morbillivirus derived by Southern blot hybridization using cloned RPV F gene as probe.

Animals↗
PubMed1988

Parameters of field inversion gel electrophoresis for the analysis of pox virus genomes.

The effects of variation in the lengths of forward and reverse pulses, voltage gradient, gel concentration and gel temperature on the mobility of DNA molecules in agarose gels during field inversion gel electrophoresis (FIGE) have been determined. A curve, which best fits the empirical data, is presented and allows the choice of pulse conditions and voltage gradient most suitable for the resolution of molecules of chosen size. The use of FIGE in the analysis and direct mapping of large virus genomes is illustrated using vaccinia virus DNA.

DNA, Viral↗
PubMed1988

Studies on the infectivity of foot-and-mouth disease virus RNA using microinjection.

Foot-and-mouth disease virus (FMDV) RNA, isolated as virion RNA from purified virus particles or as total RNA from infected cells, has been microinjected into nuclei and cytoplasms of BHK cells. When injected directly into the nucleus FMDV RNA was not infectious, whereas cytoplasmic injection resulted in a high proportion of productive infections. Infectivity microinjection assays on dilution series of various FMDV RNAs showed that both single-stranded positive sense 35S RNA and double-stranded replicative form (Rf) RNA have an infectivity close to 1 p.f.u. per molecule, although only a minor fraction of BHK cells appeared able to support plaque formation following microinjection of Rf FMDV RNA. The infectivity of Rf FMDV RNA was not sensitive to inhibition by actinomycin D. The results are discussed in relation to the high virus particle to p.f.u. ratios observed for FMDV.

Animals↗
PubMed1987

Different subfamilies of alphoid repetitive DNA are present on the human and chimpanzee homologous chromosomes 21 and 22.

The alphoid repeat DNA on chimpanzee chromosome 22 was compared with alphoid repeat DNA on its human homologue, chromosome 21. Hybridization of different alphoid probes under various conditions of stringency show that the alphoid repeats of chimpanzee chromosome 22 are not closely related to those of human chromosome 21. Sequence analysis of cloned dimer and tetramer EcoRI fragments from chimpanzee chromosome 22 confirm the low overall level of homology, but reveal the presence of several nucleotide changes which are exclusive to the chromosome 21 subfamily of human alphoid DNA. Southern blot analysis of alphoid repeat DNA on the chimpanzee X chromosome suggests this subfamily has been strongly conserved during and since the separation of chimpanzee and man although the two subfamilies can be distinguished on the basis of Taq I restriction fragments.

Animals↗
PubMed1987

Homologous subfamilies of human alphoid repetitive DNA on different nucleolus organizing chromosomes.

The organization of alphoid repeated sequences on human nucleolus-organizing (NOR) chromosomes 13, 21, and 22 has been investigated. Analysis of hybridization of alphoid DNA probes to Southern transfers of restriction enzyme-digested DNA fragments from hybrid cells containing single human chromosomes shows that chromosomes 13 and 21 share one subfamily of alphoid repeats, whereas a different subfamily may be held in common by chromosomes 13 and 22. The sequences of cloned 680-base-pair EcoRI fragments of the alphoid DNA from chromosomes 13 and 21 show that the basic unit of this subfamily is indistinguishable on each chromosome. The sequence of cloned 1020-base-pair Xba I fragments from chromosome 22 is related to, but distinguishable from, that of the 680-base-pair EcoRI alphoid subfamily of chromosomes 13 and 21. These results suggest that, at some point after they originated and were homogenized, different subfamilies of alphoid sequences must have exchanged between chromosomes 13 and 21 and separately between chromosomes 13 and 22.

Base Sequence↗
PubMed1987

The transcription of Xenopus laevis embryonic U1 snRNA genes changes when oocytes mature into eggs.

X. laevis stage VI oocytes respond differently from unfertilized eggs when injected with the genes for X. laevis embryonic U1 RNAs, xU1b1, and xU1b2. Upon maturation of oocytes into eggs, the efficiency of transcription decreases greatly and the ratio of xU1b1 to xU1b2 RNA transcription changes. Moreover, DNA replication is now required for transcription. Because of differences in the 5'-flanking regions of the two xU1b genes, xU1b2 RNA transcription predominates after injection into oocytes; in contrast, xU1b1 RNA transcription predominates after injection into unfertilized eggs. Our results also indicate that in oocytes a factor that interacts with sequences close to the coding region is limiting, whereas in eggs a factor that recognizes far-upstream sequences required for enhancer activity is limiting. Qualitatively, expression of the embryonic xU1b genes injected into eggs closely resembles that of the endogenous genes during early embryogenesis.

Animals↗
PubMed1986

Structure of bovine papillomavirus type 1 DNA in a transformed mouse cell line.

Linearized bovine papillomavirus type 1 (BPV-1) DNA was introduced into mouse C127 cells, where it recircularized and replicated as an intact monomeric, extrachromosomal circular form in the resulting transformants. These cells contained a mixture of complex high molecular weight forms that were converted to a linear form of approximately BPV-1 size upon digestion with an enzyme that cuts once within the BPV-1 genome. Further analysis of one of these cell lines revealed that these high molecular weight forms consisted of two components. One was detected on agarose gels as a diffuse smear of slow-migrating material representing linear forms that were tightly associated with host chromosomes, probably by integration. The second component was composed of discrete-sized oligomeric open and supercoiled extrachromosomal circular forms of up to approximately 48 X 10(3) base-pairs (6 tandemly linked BPV-1 genomes) in size. No catenated (interlocked) forms could be detected.

Animals↗
PubMed1986

Mechanisms of DNA sequence amplification and their evolutionary consequences.

DNA sequence amplification is a phenomenon that occurs predictably at defined stages during normal development in some organisms and has been shown to occur spontaneously, but sporadically, in a variety of cells, including mammalian cells, selected for overproduction of a gene product. Developmentally programmed gene amplification includes rDNA amplification during oögenesis in amphibia, chorion protein gene amplification in Drosophila and the chromosomal changes accompanying macronuclear formation in ciliates. Selected gene amplification is illustrated by mutant mammalian cells which have been selected in vitro or in vivo for the overproduction of a gene product. In these cells the unit of DNA that is amplified is much larger than the gene under selection, and appears to be formed by multiple recombination events, which bring together sequences not normally adjacent to each other. Often the product of amplification can be seen microscopically as aberrant chromosome forms. The vast majority of DNA amplification events occur in somatic nuclei, and thus would not have any direct effect on the evolution of a genome. However, the ability to amplify DNA in somatic cells does have consequences for the composition of the genomes of the organisms in which it can occur, and should DNA amplification occur, even sporadically, in germ-line cells the potential effect on evolution would be great.

Amphibians↗
PubMed1986

Chromosome-specific subfamilies within human alphoid repetitive DNA.

Nucleotide sequence data of about 20 X 10(3) base-pairs of the human tandemly repeated alphoid DNA are presented. The DNA sequences were determined from 45 clones containing EcoRI fragments of alphoid DNA isolated from total genomic DNA. Thirty of the clones contained a complete 340 base-pair dimer unit of the repeat. The remaining clones contained alphoid DNA with fragment lengths of 311, 296, 232, 170 and 108 base-pairs. The sequences obtained were compared with an average alphoid DNA sequence determined by Wu & Manuelidis (1980). The divergences ranged from 0.6 to 24.6% nucleotide changes for the first monomer and from 0 to 17.8% for the second monomer of the repeat. On the basis of identical nucleotide changes at corresponding positions, the individual repeat units could be shown to belong to one of several distinct subfamilies. The number of nucleotide changes defining a subfamily generally constitutes the majority of nucleotide changes found in a member of that subfamily. From an evaluation of the proportion of the total amount of alphoid DNA, which is represented by the clones studied, it is estimated that the number of subfamilies of this repeat may be equal to or exceed the number of chromosomes. The expected presence of only one or a few distinct subfamilies on individual chromosomes is supported by the study, also presented, of the nucleotide sequence of 17 cloned fragments of alphoid repetitive DNA from chromosome 7. These chromosome-specific repeats all contain the characteristic pattern of 36 common nucleotide changes that defines one of the subfamilies described. A unique restriction endonuclease (NlaIII) cleavage site present in this subfamily may be useful as a genetic marker of this chromosome. A family member of the interspersed Alu repetitive DNA was also isolated and sequenced. This Alu repeat has been inserted into the human alphoid repetitive DNA, in the same way as the insertion of an Alu repeat into the African green monkey alphoid DNA.

Base Sequence↗
PubMed1986

Comparison of methods for introducing vectors based on bovine papillomavirus-1 DNA into mammalian cells.

The intracellular structure of several vectors based on BPV-1 DNA has been analyzed following transfection into mouse C127 cells by the calcium phosphate method or, for the first time, by microinjection directly into the nucleus. It is shown that the method of introduction markedly affects the fate of a BPV-1 based vector. In general, microinjection appears to do little damage to DNA and is more likely to result in a vector replicating extrachromosomally as a monomeric structure of the same size as the input DNA. The method of selection for transformed cells, e.g., focus formation versus resistance to the neomycin analog G418, can also affect the intracellular state of the BPV-1 vector DNA. The nature of the recipient mammalian cell also influences whether a vector can replicate extrachromosomally or whether it integrates. BPV-1 based vectors, which replicated predominantly as multicopy intact extrachromosomal forms in mouse C127 cells, were always found to have integrated at low copy number in mouse LtAp20 cells.

Animals↗
PubMed1986

Duplication of a viral enhancer sequence improves the stability of a vector based on BPV-1 DNA.

Various recombinant constructions involving bovine papillomavirus type 1 (BPV-1) DNA and bacterial plasmids have been tested for their ability to transform mouse C127 cells and replicate as intact extrachromosomal monomeric structures. When BPV-1 DNA was linked to pBR328, pAT153 or derivatives of these plasmids lacking the 344 bp HindIII-BamHI fragment or another small segment, the resulting vectors replicated in C127 cells as high molecular weight structures and, in some cases, deleted extrachromosomal forms. The sequences which became deleted were generally the non-BPV-1 sequences. Duplication of the 3' distal enhancer sequence of BPV-1 DNA in one of the vectors increased its stability upon introduction into C127 cells, but some deleted and high molecular weight forms were still observed.

Animals↗
PubMed1984

Chromosomal changes associated with changes in development.

In the past there has been a tendency to dwell on aspects of chromosomes which stress constancy of structure, number and content; even to the extent of dismissing exceptions as 'aberrations' or 'oddities'. It is now becoming clear that genomes can be quite plastic, and that structural changes to chromosomes are an important and often necessary part of normal differentiation and development. Elimination of whole chromosome sets or defined portions of genomes is not uncommon and selective gene amplification has been shown to be part of normal development in both protozoa and higher organisms. Chromosomal rearrangements are now a well-documented feature of normal development of, for example, B- and T-lymphocytes and trypanosomes. Transposable elements, whose mobility may not be part of normal developmental processes, can have marked effects on development if their transposition takes them to developmentally important genes. This article reviews some of the structural changes that occur during normal development, and discusses some of the consequences for development when the mechanisms which bring about these rearrangements go wrong.

Animals↗
PubMed1983

Gene amplification in methotrexate-resistant mouse cells. V. Intact amplified units can be transferred to and amplified in methotrexate-sensitive mouse L cells.

Wild-type mouse LtAp20 cells were treated with calcium phosphate-precipitated DNA or chromosomes from two highly Methotrexate (MTX)-resistant mouse lymphoma cell lines--EL4/8 and EL4/11. Transfections with purified MTX-resistant DNA produced colonies of LtAp20 cells resistant to 3 X 10(-8) M MTX, at about eight times the frequency with which resistant colonies arose in control transfections. DNA transfectants contained multiple copies of the dihydrofolate reductase (dhfr) gene, but other sequences characteristic of the donor DNA could not be detected. Transfections using isolated chromosomes were twice as efficient as those using purified DNA. Unlike DNA transfectants, over 90% of all chromosome transfectants took up large stretches of donor DNA intact and contained DNA sequences characteristic of donor DNA. Of chromosome transfectants selected for resistance to high levels of MTX (1 mM), 70% amplified a unit of DNA which was indistinguishable from that present in the donor cell. The results showed that large fragments of chromosomes (as opposed to purified DNA) can be taken up to recipient cells without detectable alteration to the fine structure of the DNA they contain. The results also support the notion that all amplified units within a MTX-resistant cell have the same overall complex DNA structure.

Adenine Phosphoribosyltransferase↗
PubMed1982

Gene amplification in methotrexate-resistant mouse cells. IV. Different DNA sequences are amplified in different resistant lines.

DNA was purified from double minutes isolated from MTX-resistant EL4/8 mouse lymphoma cells, digested to completion with Bam H1 restriction endonuclease and cloned in lambda-1059. The properties of the library suggest that the DNA from which it was made was not detectably contaminated with non-dm chromosome material, and that the library is essentially complete for sequences contained in Bam H1 restriction fragments between 9 and 19 kb. The inserts of some selected lambda-recombinants were subcloned in pBR328 or pAT153 to separate sequences of differing repetition frequency. Clones representative of different classes of sequences were used as probes to Southern transfers of Bam H1 digested total nuclear DNAs of various MTX-resistant cell lines. The results clearly show that the amplified unit of each cell line has a unique structure, and that different amplified units differ widely in their sequence composition.

Animals↗
PubMed1982

Molecular biology and the clinician.

Molecular biology has uncovered informational processes which broadly apply in medicine. The points of attack range from diagnosis at the most fundamental level of information flow to the production of therapeutic agents. This is promoting movement from serendipitous strategies of advance to those of predictive rational clinical design. Specific examples show how three principal types of molecular probe are benefiting a wide range of clinical disciplines and thereby blurring interdisciplinary boundaries. There are signs that current medical training is not making the best use of contemporary molecular biology. The identification of areas in which molecular biology can be usefully applied to medicine requires medical practitioners to be aware of its diagnostic and prognostic potential. Equally, advances are delayed by gaps between laboratory workers and their clinical colleagues. The dilemma is that by creating a specialist discipline of clinical molecular biology we risk it becoming isolated and delay the impact of molecular biology in medicine as a whole. An alternative is to find a means for raising molecular consciousness throughout all disciplines in medicine.

Cloning, Molecular↗