Studies on the infectivity and cytopathology of epstein‐barr virus in human lymphoblastoid cells

Durr, Frederick E.; Monroe, James H.; Schmitter, R.; Traul, Karl A.; Hirshaut, Yashar

doi:10.1002/ijc.2910060315

Cited by 85 publications

(28 citation statements)

References 16 publications

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“…Cell lines containing EBV plasmids were created from CV-1 by transfection of 10 ,ug of plasmid DNA by using calcium phosphate coprecipitation which was followed by selection in medium containing hygromycin at a concentration of 200 jig/ml. NC-37 is a human lymphoid cell line (6).…”

Section: Methodsmentioning

confidence: 99%

Autonomous DNA replication in human cells is affected by the size and the source of the DNA.

et al. 1991

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We previously developed short-term and long-term assays for autonomous replication of DNA in human cells. This study addresses the requirements for replication in these assays. Sixty-two random human genomic fragments ranging in size from 1 to 21 kb were cloned in a prokaryotic vector and tested for their replication ability in the short-term assay. We found a positive correlation between replication strength and fragment length, indicating that large size is favored for efficient autonomous replication in human cells. All large fragments replicated efficiently, suggesting that signals which can direct the initiation of DNA replication in human cells are either very abundant or have a low degree of sequence specificity. Similar results were obtained in the long-term assay. We also used the same assays to test in human cells a random series of fragments derived from Escherichia coli chromosomal DNA. The bacterial fragments supported replication less efficiently than the human fragments in the short-term and long-term assays. This result suggests that while the sequence signals involved in replication in human cells are found frequently in human DNA, they are uncommon in bacterial DNA.In this study we investigated the genetic requirements for origin of replication function in human cells. The study utilizes a system we have developed that allows us to test fragments of DNA for autonomous replication when introduced into human tissue culture cells in both long-and short-term assays. This system should ultimately permit us to deduce the DNA sequence requirements for autonomous replication. It is likely that this information will be relevant for chromosomal replication as well.Over the course of many cell divisions, extrachromosomal vectors are lost from the nuclei of mammalian cells unless they have a means of nuclear retention. The long-term replication assay that we developed overcomes the problem of plasmid loss by using a replication-defective vector derived from Epstein-Barr virus (EBV) that permits linked sequences to be retained in human cells (22). When random human DNA was cloned into this vector and introduced into human cells, we found that a large number of fragments could confer replication ability on the vector in a long-term assay. The resulting plasmids were stably maintained for at least 2 months in human cells.The fragments could also provide for replication in our short-term assay (22). In this assay, the fragments are moved to a prokaryotic vector that lacks all viral sequences, and DNA is harvested 4 days after transfection, since the plasmids have no means of nuclear retention. These experiments indicated that the human DNA fragments that we had isolated contained sequences involved in the initiation of DNA replication in human cells. Subsequent studies have shown that, like the chromosomes, these plasmids undergo replication once per cell cycle in human cells (unpublished data). This characteristic makes the autonomously replicating system a reasonable model for chromosomal replication.The hu...

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Section: Methodsmentioning

confidence: 99%

Autonomous DNA replication in human cells is affected by the size and the source of the DNA.

et al. 1991

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“…Patients-.Blood samples were obtained from hospitalized and ambulatory patients in several local hospitals (Table J) (Henderson, 1977) (Hinuma et al, 1967), Daudi (Klein et al, 1968) and NC-37 (C-6) (Durr et al, 1970) (Lozzio & Lozzio, 1973 Separation of mononuclear cells.-Mononuclear cells were separated from fresh heparinized peripheral blood or marrow aspirates obtained from patients and healthy donors by Ficoll-Conray density gradient centrifugation. They were washed x 3 with Eagle's minimum essential medium and resuspended with RPMI-1640 (both from Nissui Seiyaku Co. Ltd, Tokyo) supplemented with 10% foetal calf serum (Grand Island Biological Co., Grand Island, N.Y.) at a concentration of 106 cells/ml.…”

Section: Methodsmentioning

confidence: 99%

Fluorescence polarization with FDA in leukaemic cells: a clear difference between myelogenous and lymphocytic origins

Tsuda¹,

Maeda²,

Kishimoto³

1981

Br J Cancer

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Summary.-Intracellular fluorescence polarization (IFP) values of normal human lymphocytes and leukaemic cells from newly diagnosed patients were determined from fluorescence polarization using fluorescein diacetate (FDA). Thirty healthy donors and 40 patients with various types of leukaemia (20 myelogenous and 20 lymphocytic) were included in the present studies. The result was that myeloid cells had about twice the polarization value of lymphocytic cells. The use of FDA for the determination of IFP appears to be useful for differential diagnosis, at least between acute myelogenous and lymphocytic leukaemias. These 2 types of leukaemia also showed a pronounced difference in fluorescence intensity when treated with FDA, perhaps owing to a difference in uptake velocity. The previously described membrane microviscosity using 1,6-diphenyl-1,3,5-hexatriene (DPH), however, did not show such a difference between these 2 leukaemias.The fluorescein-binding protein(s) was also investigated in order to clarify its effect on IFP, but there seemed little evidence for the existence of any such dyebinding protein(s). The advantages of the present method, using FDA, reside in its simplicity, rapidity and considerable sensitivity, requiring a small sample of blood usually <5 ml.

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“…Other HTLV-negative cell lines used were as follows. T cell lines; RPMI 8402 (Strivastava et al, 1975), (Strivastava & Minowada, 1973), and CCRF-CEM (Foley et al, 1965): B cell lines; Namalwa (Klein et al, 1972), CESS (Muraguchi et al, 1981), NC 37 (Durr et al, 1970), and BALL-1 (Miyoshi et al, 1977); myeloma cell lines; (Burk et al, 1978) and RPMI 8226 (Matsuoka et al, 1967). They were maintained at 37°C in RPMI 1640 (M.A.…”

Section: Cell Linesmentioning

confidence: 99%

Production of plasminogen activators by human T-cell leukaemia virus-transformed human T cell lines

Hinuma¹,

Honda²,

Tsukamoto³

et al. 1985

Br J Cancer

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Studies on the infectivity and cytopathology of epstein‐barr virus in human lymphoblastoid cells

Abstract: The infectivity of EB virus recovered from cultures of P-3J and the H R l K

Cited by 85 publications

References 16 publications

Autonomous DNA replication in human cells is affected by the size and the source of the DNA.

Autonomous DNA replication in human cells is affected by the size and the source of the DNA.

Fluorescence polarization with FDA in leukaemic cells: a clear difference between myelogenous and lymphocytic origins

Production of plasminogen activators by human T-cell leukaemia virus-transformed human T cell lines

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