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Biomedical subjects

L Manuelidis

Publications and source records attributed to L Manuelidis.

At least 91 records · Page 5Linked to original sources
PubMed1982

Nucleotide sequence definition of a major human repeated DNA, the Hind III 1.9 kb family.

A human Hind III 1.9 kb repeated DNA fragment was isolated and cloned in pBR322. A cloned member that hybridized predominantly to the 1.9 kb Hind III band in a digest of whole human DNA was chosen for sequencing. It is an 1894 bp fragment that shows no significant internal repeats. Few pCG residues are observed in the sequence and there are numerous stop codons. Detailed sequence comparisons confirm this is a novel class of repeats that is not related to previously characterized human satellite DNAs or Alu sequences. At least a portion of the sequence described is conserved in evolution.

Animals↗
PubMed1982

Genomic representation of the Hind II 1.9 kb repeated DNA.

The genomic representation and organization of sequences homologous to a cloned Hind III 1.9 kb repeated DNA fragment were studied. Approximately 80% of homologous repeated DNA was contained in a genomic Hind III cleavage band of 1.9 kb. Double digestion studies indicated that the genomic family, in the majority, followed the arrangement of the sequenced clone, with minor restriction cleavage variations compatible with a few base changes. Common restriction sites external to the 1.9 kb sequence were mapped, and hybridization of segments of the cloned sequence indicated the 1.9 kb DNA was itself not tandemly repeated. Kpn I bands which were homologous to the sequence contained specific regions of the repeat, and the molecular weight of these larger fragments could be simply explained. Mapping of common external restriction sites indicated that in some but not all cases the repeat could be organized in larger defined blocks of greater than or equal to 5.5 kb. In some instances, flanking regions adjacent to the repeat may contain common DNA elements such as other repeated DNA sequences, or possibly rearranged segments of the 1.9 kb sequence. It is suggested that although the 1.9 kb sequence is not strictly contiguous, at least some of these repeated sequences in the human genome are arranged in clustered or intercalary arrays. A region of the 1.9 kb sequence hybridized to a mouse repeated DNA, indicating homology beyond the primates.

Base Composition↗
PubMed1982

High-resolution mapping of satellite DNA using biotin-labeled DNA probes.

We have developed a novel method for high resolution mapping of specific DNA sequences after in situ hybridization. DNA probes, labeled with biotin-nucleotides in conventional nick-translation reactions, are hybridized to cytological preparations and detected with affinity-purified rabbit antibiotin antibodies followed by antibodies to rabbit IgG that are conjugated to fluorescent or enzymatic reagents. Using peroxidase labeled anti-rabbit IgG, we are able to detect and localize specific sequences at both the light and electron microscopic levels. Initial studies were done with repeated DNA sequences previously mapped by light microscope autoradiography to assess the fidelity and resolution of this method. An analysis using biotin-labeled mouse satellite DNA is presented here.

Animals↗
PubMed1980

Novel classes of mouse repeated DNAs.

Mouse DNA cleaved with Eco R11 (bst NI) displays two prominent restriction bands of 1.5 and 1.7 kb in agarose gels stained with ethidium bromide. These constitute novel subsets of repeated DNA in the mouse. Sequential Hoechst 33258-CsCl gradient fractionation of mouse DNA, yielding more GC rich main band DNA, and AT rich satellite DNA, revealed that both these fragments copurified with GC rich main band DNA. They were not detected in purified satellite preparations. Together these restriction bands constituted larger than or equal to 0.2% of main band DNAs. Hybridization of 32p labelled satellite DNA to blots of Eco R11 restricted mouse DNA showed positive hybridization only to smaller satellite restriction fragments, indicating satellite DNA had little or no homology with either the 1.5 or 1.7 kb fragments. The 1.5 and 1.7 kb fragments were isolated from gels and labelled with 32p by nick translation. Using a series of restriction endonucleases each of these two fragments showed different cleavage patterns. Filter hybridization confirmed that these two fragments were distinct subsets as they did not cross hybridize with each other. They also did not hybridize to other more minor repeated non-satellite DNA bands noted in ethidium bromide stained gels. Neither of them could be assigned to ribosomal genes as they did not hybridize to 32p kinase labelled 18S and 28S RNA. Isolation of DNA from male and female mice showed comparable amounts of both the 1.5 and 1.7 kb fragments. Thus neither was Y chromosome specific. From restriction patterns, and preliminary chromosome hybridization studies, these fragments are thought to represent "interspersed" repeated sequences rather than very long tandem (satellite like) centromeric arrays. The relationship between these repeated sequence subsets, their evolution and detailed organization, and their representation in different mouse species, remain to be determined.

Animals↗
PubMed1980

Soft x-ray lithographic studies of interphase chromosomes.

Soft x-ray absorption lithograph patterns of purified interphase human nuclei and chromosome arrays, imaged on PMMA resists, were examined by scanning EM. The patterns obtained were compared to those utilizing more conventional sources, including transmission EM, scanning EM, high voltage EM, and various light microscopic techniques. The x-ray resist images revealed orderly arrays of absorption profiles in the 3-dimensional specimen with both mild and more extensive developments of the resist. Dense chromatin at the edge of interphase nuclei revealed aligned periodic peaks on the order of 2200 A diameter, with substructure. The periodicity and alignment of interphase chromosomes were entirely consistent with birefringent data on nuclei indicating a high degree of 3-dimensional order. This degree of 3-dimensional order was observed in nuclei containing essentially DNA and histones with only very few other minor (probably structural) proteins. Sonication and nuclease treatment to disperse interphase chromosomes revealed similar absorption periodicities in individual chromosome fibers. Analysis of x-ray absorption profiles thus appears to offer significant new insights into the ordered structure of these defined biological specimens.

Cell Cycle↗
PubMed1979

Conservation of repeated DNA sequences in aneuploid human tumor cells.

A series of human neuroectodermal tumors, all containing more than the normal diploid DNA, and each with its own distinct chromosome mode, were studied using restriction enzyme cleavage and specific DNA sequence hybridization. Methods described were quite sensitive and quantitative and as few as 40 molecules with a given restriction site were reproducibly detected in total nuclear DNA. Analysis of several fluorescent gel bands associated with different chromosomal domains revealed no changes between any of the tumor and normal cells. Specific probe hybridization, using purified complex repeating sequences, indicated fidelity of base sequence, as well as preservation of the relative amounts of each of a number of minor related multimers in both the tumor and normal cells. Centromeric regions containing arrays of such sequences may be maintained in these tumor cells and furthermore it is possible that some of these cells are polyploid with respect to DNA sequences, rather than aneuploid as their chromosome profiles suggest.

Aneuploidy↗
PubMed1978

Viremia in experimental Creutzfeldt-Jakob disease.

Inoculation of the buffy coat of blood from guinea pigs infected with Creutzfeldt-Jakob disease resulted in passage of this disease to recipient animals. This demonstrates that there is a viremia in experimental Creutzfeldt-Jakob disease. These findings suggest that the hematogenous route may be implicated in the human infection and that the disease may possibly be transmitted by blood transfusions.

Animals↗
PubMed1978

Complex and simple sequences in human repeated DNAs.

Highly repeated human DNA sequences were isolated by isopycnic centrifugation, or were eluted from gels after restriction enzyme cleavage. High molecular weight DNA peaks separable from the bulk of the DNA in a variety of gradients were shown to consist of very simple sequences characteristic of simple satellite DNAs; DNA fingerprint studies indicated each of these peaks could consist of tandem repeats of a specific oligonucleotide sequence as low as 10 base pairs (bp) long. All the gradient peaks could be assigned to one of two sequence groups and several "different" buoyant density peaks revealed the same sequence.--Restriction fragment multimers did not share common sequences with the satellite DNAs as judged by hybridization data. They could not be separated by isopycnic centrifugation. Furthermore these highly repeated DNAs were more complex in sequence and more variable than the satellites. Even the smallest (50 bp) fragments by depurination and other direct sequencing methods were shown to be more complex than the high molecular weight satellite peaks.--The idea that subsets of repeated DNAs may be defined by sequence complexity, possibly with discrete or separable functions, is proposed.

Base Sequence↗
PubMed1978

Chromosomal localization of complex and simple repeated human DNAs.

Complex repeating restriction multimers and a simple AT rich satellite isolated with Hoechst 33258 ( less than or equal to 0.5% of the human genome) were localized by in situ hybridization to human chromosomes. The complex repeats were clustered at the centromeres, consonant with their integration in tandem arrays at these loci; these sequences were very prominent on chromosomes 7, 10 and 19, sites not previously identified with any specific human repeated sequence. The Hoechst simple satellite labelled predominantly the long arms of the Y chromosome. Although this simple satellite and the complex restriction multimers did not hybridize with each other, and did not contain detectable ribosomal sequences, both isolates additionally labelled the nucleolus organizing regions (NORs) of acrocentric chromosomes.--The possible relationship of complex and simple repeated DNAs, and their assignment to specific chromosomal domains, is discussed.

Base Sequence↗
PubMed1978

Interspecies transmission of Creutzfeldt-Jakob disease to Syrian hamsters with reference to clinical syndromes and strains of agent.

Experimental Creutzfeldt-Jakob disease was serially transmitted from guinea pigs to Syrian hamsters with 100% incidence, morbidity, and mortality. All animals developed a subacute spongiform virus encephalopathy with neuronal destruction and concomitant astrocytic changes. In the first passage three different clinical syndromes were recorded, each with widely variant incubation times; these results suggested there may be different strains of the Creutzfeldt-Jakob agent, some of which may be partially separated when the agent is passaged from one species to another. Accumulations of neurofilaments in neuronal perikarya and processes and increased lipofuscin were suggestive of changes seen in senility and aging.

Animals↗