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At least 19 recordsLinked to original sources
PubMed1978

Episomal viral DNA in herpesvirus saimiri-transformed lymphoid cell lines.

Structural analysis of episomal viral genomes from two herpesvirus saimiri (HVS)-transformed tumour cell lines (No. 1670 and 70N2) showed that both types of episomes have a higher molecular weight than linear virion DNA. The arrangement of unique (L) and repetitive (H) DNA in No. 1670 episomes was studied by partial denaturation mapping. Part of the L-sequences present in linear virion DNA was found to be missing, part was found to be duplicated in the episomes. The episomal L-DNA regions were correlated with the known physical gene maps of linear HVS DNA.

Animals↗
PubMed1976

[Study of the penicillinase of the plague microbe, the synthesis of which is determined by the R-episome].

Penicillinase produced by a culture of the plague microbe with episomic resistance to benzylpenicillin was isolated and purified. Comparative investigation of the substrate spectrum, Michaelis constant and antigenic properties of the enzymes from the plague and dysentery microbes (R-factor donor) showed that transfer of the R-episome to the new host did not affect the investigated properties of penicillinase the synthesis of which was controlled by this episome.

Culture Media↗
PubMed1976

Regulation of enzyme formation in Klebsiella aerogenes by episomal glutamine synthetase of Escherichia coli.

We studied the physiology of cells of Klebsiella aerogenes containing the structural gene for glutamine synthetase (glnA) of Escherichia coli on an episome. The E. coli glutamine synthetase functioned in cells of K. aerogenes in a manner similar to that of the K. aerogenes enzyme: it allowed the level of histidase to increase and that of glutamate dehydrogenase to decrease during nitrogen-limited growth. The phenotype of mutations in the glnA site was restored to normal by the introduction of the episomal glnA+ gene. These results are consistent with the hypothesis that glutamine synthetase regulates the function of its own structural gene.

Ammonia-Lyases↗
PubMed1975

T7 protein synthesis in F-factor-containing cells: evidence for an episomally induced impairment of translation and relation to an alteration in membrane permeability.

T7 infection of F-factor-containing PIFA+, B+ cells is abortive. In spite of the presence of mRNA for all three classes of T7 proteins, only the earliest of the T7 proteins are synthesized. A crucial question is whether the failure of T7 to develop in PIFA+, B+ cells is the result of an inability to translate the late classes of T7 mRNA or, as has been recently suggested (Britton, and Haselkorn, 1975; Condit, 1975), whether it is the result of a more generalized alteration in membrane permeability. We have examined the effects of the wild-type PIFA+, B+ spisome and two sipsomal mutations (pifA- and pifB-) on in vitro translation and membrane permeability. In vivo the episomal mutations allow partial or complete T7 development to occur. We demonstrate that cell-free protein-synthesizing systems from T7-infected PIFA+, B+ cells show a three- to fivefold decrease in the rate of translation of both natural and synthetic mRNA. In addition, ribosomes from T7-infected PIFA+, B+ cells are defective in their ability to bind Fmet tRNAf in response to natural mRNA. By contrast, cell-free extracts from T7-infected pifA-(PIFA-, B+) celld retain the ability to bind Fmet defective T7-infected PIFA+, B+ rigosomes can be restored to full activity by a trypsin-sensitive fraction from uninfected PIFA+, B+ or T7-infected PIFA-, B+ cells. Despite the differences in translational capacity of these extracts, both T7-infected PIFA+, B+ and PIFA-, B+ cells display the same permeability lesions as measured by the loss of ATP from the cells into the supernatant. Mutation of the episome of pifB- prevents the loss of ATP from the cells after T7 infection.

Adenosine Triphosphate↗
PubMed2026

Generation of Aneuploid Human Induced Pluripotent Stem Cells from Primary Amniotic Fluid Cells via Episomal Plasmid Electroporation.

The generation of patient-specific induced pluripotent stem cells (iPSCs) from amniotic fluid cells (AFCs) carrying defined chromosomal aneuploidies provides a powerful platform for modeling genetic disorders. However, establishing a reliable and reproducible reprogramming pipeline for aneuploid AFCs remains technically challenging due to the intrinsic genomic instability and variable proliferative capacity of these cells. Here, we present a comprehensive, non-integrating method for generating aneuploid human iPSCs from primary AFCs using episomal plasmid electroporation. This protocol details the complete workflow, encompassing cell thawing and expansion with a gradual media adaptation strategy, optimized plasmid delivery via electroporation system, sequential post-electroporation culture with mesenchymal-to-epithelial transition (MET)-directed media changes, and mechanical colony picking based on defined morphological criteria. We further describe validation procedures, including immunofluorescence staining for core pluripotency markers, G-banding karyotype analysis to confirm aneuploid karyotype maintenance, and PCR-based episomal vector clearance verification. This feeder-free, integration-free protocol yields aneuploid iPSC lines suitable for disease modeling, drug screening, and studies of chromosome biology.

Humans↗
PubMed1975

[Study of the mechanisms of levomycetin inactivation by the palgue causative agent and Escherichia coli with episomal and chromosomal resistance. The enzymatic acetylation of levomycetin].

One of the mechanisms of levomycetin inactivation, i.e. enzymatic acetylation of the plague causative agent and Coli bacteria with chromosomic and episomic type of resistance was studied. It was shown that resistance to levomycetin in the recombinants of the plague microbe and Coli bacteria stipulated by one and the same R-factor was associated with their capacity for the antibiotic inactivation mainly with the help of levomycetinacetyltransferase. At the same time the enzyme activity in the mutants was very low and in the cells of Y. pestis and E. coli sensitive to the antibiotic it was absent. Levomycetinacetyltransferase is to somee extent soluble, still a singificant amount of it is connected with the structures of the bacterial cells and in particular with the cytoplasmic membranes. Comparison of the activity of levomycetinacetyltransferase as dependent on the incubation time, substrate concentration, pH and thermal treatment provided determination of quantitative differences in the properties of the enzyme of Y. petis and E. coli with episomic multiple drug resistance. However, under the experimental conditions transfer of R-factor from one host to the other induced no qualitative changes in the mechanism of levomycetin inactivation.

Acetylation↗
PubMed1976

Circular DNA of a yeast episome with two inverted repeats: structural analysis by a restriction enzyme and electron microscopy.

Small circular DNA molecules from genetically characterized clones of Saccharomyces cerevisiae have been studied by restriction endonuclease analysis and electron microscopy. The circular monomers (6000 bases) are shown to contain two inverted repeats of the same sequence (600 bases) situated opposite each other along the perimeter. Four endonuclease EcoRI fragments are obtained in 1:1:1:1 stoichiometry, and their sum gives a length of about 12,000 bases. The two large fragments and the two small ones differ from each other by 200 bases. We propose a model for the structure of the monomer molecule. Two classes of monomers can be generated by intramolecular recombinations within inverted repeats; they differ by the relative orientation of nonrepeated segments. The structure of dimers as predicted by the model is verified by self-renaturation of single-stranded circles. Inverted repeats in circular molecules may be related to the insertion release faculty of II episome in the chromosomes.

Base Sequence↗
PubMed1979

Persistent BK papovavirus infection of transformed human fetal brain cells. I. Episomal viral DNA in cloned lines deficient in T-antigen expression.

After infection of permissive human fetal brain cells by BK human papovavirus (BKV), the vast majority of the cells were killed by the virus, but rare survivors were recovered after frequent medium changes. These surviving cells grew and formed visible colonies after 5 to 6 weeks and were thereafter established as permanent cell lines. These cells, designated as BK-HFB cells, were persistently infected and shed BKV. Morphologically, they were small polygonal cells and had transformed growth properties. Their plating efficiency on solid substrates or in semisolid medium was high, and they were tumorigenic in athymic nude mice. Cloning experiments in medium containing BKV antiserum revealed that BKV did not persist in the cultures in a simple carrier state. All cloned cell lines were initially T-antigen negative and virus-free. However, every clone began to release BKV and again became persistently infected within 3 weeks after removal of BKV antiserum. After rigorous antibody treatment, four of seven clones still released virus spontaneously upon removal of antiserum; three clones have remained virus-free and are apparently cured. Although these cloned cell lines are T- and V-antigen negative when grown in antiserum-containing medium, they retain "free" or episomal BKV genomes; integrated viral DNA was not detected in any of the clones. These free genomes are indistinguishable from prototype BKV DNA and are found in much larger amounts in virus-shedding cell lines.

Antigens, Neoplasm↗
PubMed1977

Transfer of episome F'lac+ and chromosomal trp+ genes from Erwinia amylovora to Salmonella typhimurium.

The F'lac+ episome of Escherichia coli origin was transferred by conjugation with frequencies of 10(-7) to 10(-5) from Erwinia amylovora to 14 out of 15 Salmonella typhimurium trp female parents. The chromosomal trp+ genes were transferred with frequencies of 10(-7) to 10(-6) only to one trpB and 2 trpD female parents, which have a point mutation in the 2nd and fourth structural genes, respectively, of the tryptophan operon. The transferred male trp+ genes became integrated at the selected sites of the S. tryphimurium chromosome. The resulting Trp+ hybrids were phenotypically stable, lacked a cryptic trp allele selected against in the female parent, had high genetic homology values in the tryptophan region, and showed biochemical reactions and pathogenicity typical of S. typhimurium.

Animals↗
PubMed1976

[Formation of F'-episomes in Flexner shigellae].

Genetic characteristics of Shigella flexneri virulent donor strain 3s, isolated in the laboratory, are studied. The data obtained showed that Sh. flexneri 3s 6832 is of F'protype it did not transmit even closely located chromosome markers, except pro+. The transition of pro+ marker was reproduced in recA- strain. F'pro plasmid was eliminated with a frequency similar to that of F'lac factor in the control strain of Escherichia coli. The study of virulence by means of keratoconjunktival test revealed its retaining in clones which lost F'pro plasmid. Probably, KCP-A gene retained in the chromosome of F'pro strain. Crossing with proA- and procC- C. coli strains revealed a small length of F'pro episome, enly ProA+ hybrids being obtained. A complex mechanism of F'system formation in interspecies genetic recombination between E. coli K-12 and Sh. flexneri 3s is demonstrated.

Extrachromosomal Inheritance↗
PubMed1975

Initiation of DNA replication in Escherichia coli. II. Effect of rifampicin on the resumption of replication of F episome and chromosome upon the returning of dna mutants from a non-permissive to a permissive temperature.

When E. coli F+ cells carrying the dna-167 or dnaC2 mutation, which causes the temperature-sensitive initiation of DNA replication, are exposed to a non-permissive temperature to stop the replication of chromosome and F factor, and then transferred back to a permissive temperature with the addition of chloramphenicol, one round of the chromosomal replication occurs, but further replication is inhibited. Under these conditions, F DNA replicates coincidentally with the initiation of the chromosomal replication in both strains. When rifampicin is added to the cells upon lowering of the temperature, the chromosome can not replicate in the F+ dna-167 strain, but can do so in the F+ dnaC2 strain. F DNA can replicate in both of the mutant strains under these conditions.

Chloramphenicol↗
PubMed1977

Determinant for multiple drug resistance possessing features of a mitochondrial episome in Saccharomyces cerevisiae.

A mutation for multiple resistance to tetracycline, cycloheximide and oligomycin appears to be followed by reconstruction of the mitochondrial genome resulting in the formation of independent nucleotide sequences that determine different resistant phenotypes. Heterozygotes for the cross resistance factor lack locus T responsible for relation tetracycline which comes from the alpha-parent. The nuclear recessive gene-suppresor i induces deletion of the whole determinant for multiple resistance. The loss of mt-DNA on ethidium bromide treatment does not lead to the loss of this determinant which remains in the cells either in an active or in a passive state.

Alleles↗
PubMed1977

Episomal viral DNA in a Herpesvirus saimiri-transformed lymphoid cell line.

The lymphoid cell line #1670 has been derived from the infiltrated spleen of a tumor-bearing marmoset monkey infected with Herpesvirus saimiri. The cells contain both types of H. saimiri DNA, unique light (L-) DNA (36% cytosine plus guanine) and repetitive heavy (H-) DNA (71% cytosine plus guanine), without producing infectious virus. Viral DNA was found to persist in these cells as nonintegrated circular DNA molecules. Closed circular superhelical viral DNA molecules were isolated by three subsequent centrifugation steps: (i) isopycnic centrifugation in CsCl, (ii) sedimentation through glycerol gradients, and (iii) equilibrium centrifugation in CsCl-ethidium bromide. The isolated circles had a molecular weight of 131.5 +/- 3.6 x 10(6). This is significantly higher than the molecular weight of linear DNA molecules isolated from purified H. saimiri virions (about 100 x 10(6)). Partial denaturation mapping of circular molecules from #1670 lymphoid cells showed uniform arrangement of H- and L-DNA sequences in all circles. All denatured molecules contained two L-DNA regions (molecular weights of 54.0 +/- 1.8 x 10(6) and 31.5 +/- 1.3 x 10(6)) and two H-DNA regions (molecular weight of 25.6 +/- 1.9 x 10(6) and 20.0 +/- 0.8 x 10(6)) of constant length. Maps of both L-regions suggested that the sequences of the shorter L-DNA region were a subset of those of the longer region. The sequences of both L-regions had the same orientation. Circular molecules from H. saimiri-transformed lymphoid cell line #1670 appeared to represent defective genomes, containing only 75% of the genetic information present in L-DNA of H. saimiri virions.

Animals↗
PubMed1975

Isolation of a hybrid F' factor-carrying Escherichia coli lactose region and Salmonella typhimurium histidine region, F42-400 (F' ts114 lac+, his+): its partial characterization and behavior in Salmonella typhimurium.

Episome F' ts114 lac+ (F42-114) was transferred into Salmonella typhimurium carrying an F'his+ (FS400) episome, and fused episome F' ts114 lac+, his+ (F42-400) was obtained. Episome F42-400 could be transferred to S. typhimurium, Escherichia coli and Klebsiella pneumoniae. Identification of the episome was based on: (i) temperature sensitivity of the Lac+ and His+ phenotypes; (ii) the fact that F- segregants, obtained after temperature curing or acridine orange curing, were simultaneously Lac- and His-; and (iii) linkage of lac+ with his+ in episomal transfers to E. coli and S. typhimurium. The frequency of episome transfer was influenced by the genotype of the donor. Plasmid LT2, prevalent in S. typhimurium LT2 strains, was suggested to be responsible for the low fertility of S. typhimurium donors. Episome F42-400 was capable of chromosome mobilization, and the extent of chromosome mobilization was not influenced by the presence or absence of the histidine region on the donor chromosome. Growth in a defined medium with acridine orange was able to cure F42-400. The frequency of curing was increased (the frequency of His+ cells was 0.0001%) if the cells were grown at 40 C in the presence of acridine orange. Selection for temperature-resistant Lac+, His+ derivatives in a strain without histidine deletion yielded Hfr strains. However, similar and stronger selections in strains without the chromosomal histidine region failed to yield Hfr strains. Our inability to obtain Hfr's in strains without the chromosomal histidine region was explained by assuming that the episome F42-400 has lost the F sites involved in integration into the S. typhimurium chromosome.

Acridines↗