Circular DNA's from HeLa cell nuclei and mitochondria

An electron microscopic observation was made on the DNA's extracted from purified HeLa cell nuclei, mitochondria, and the whole cell, and fractionated by ethidium bromide-cesium chloride density gradient method or sucrose density gradient method. Nuclear DNA presents mainly long linear DNA derived from fragmented chromosomal DNA. In addition to this, the existence of small circular DNA molecules measuring 0.32 -1.78 &#956;, was confirmed. Mitochondrial DNA was mainly circular DNA, which measured 4.87 &#956; in the mean value of the contour lengths in the highest frequency group, and small circular DNA molecules, measuring 0.3-1.01 &#956; in contour length, were also found in an extremely low frequency.</p

Relationship between molecular length and biological activity of SV40 DNA

The correlation of infectivity to the length of SV 40 DNA by dilute and undiluted passage was described. DNA was extracted from purified virions by Marmur's method, and propagation of virus was done using VERO cells. The infectivity of SV 40 (simian virus 40) ran parallel with the length of its DNA. Undiluted passage caused shortening of DNA and decrease in infectivity, but when these undiluted group was passaged dilutely, length of DNA approached the original length and the infectivity recovered. In undiluted passage group small circular DNAs under 1.0,11_, so far not reported in the SV 40 DNA, were found in low frequency. Replicating form and dimers were also noted from virions and the nuclei of SV 40 infected VERO cells.</p

Finestructure and template activities of DNA-histone complexes reconstituted in the presence and absence of urea.

Several DNA-histone complexes were reconstituted in the presence and absence of urea. The fiber size of DNA-histone H1 complex was about 20 A in width with knobs 100 to 250 A in diameter interspersed at an average interval of about 1,100 A. H1-was associated with DNA segments corresponding to a DNA size of fewer than 100 base pairs. DNA-histones H2A, H2B, H3 and H4 complex consisted of globular subunits 100 to 150 A in diameter alternating with thin strands, like beads on a string. DNA-whole histones complex was 200 to 250 A in width and had a condensed configuration. The nuclease digestion pattern of the complexes containing histones H2A, H2B, H3 and H4 was regular, similar to that of chromatin, and was disrupted by urea. The complex containing H1 was inactive for in vitro RNA synthesis by escherichia coli RNA polymerase, whereas the other complexes were active. The complexes reconstituted in the absence of urea had template activities slightly less than in the presence of urea

Thermal denaturation and template activities of reconstituted DNA-histone complexes

Reconstituted complexes of DNA with histone were prepared by salt-and-urea step gradient dialysis. The DNA was complexed with histone H1, with the combination of the other four histones H2A, H2B, H3 and H4, and with whole histones. These DNA-histone complexes were purified by Bio-Gel column chromatography, and the weight ratio of histone-to-DNA was determined in each complex. The thermal denaturation profile and nuclease digestion pattern of DNA-histone H2A, H2B, H3 and H4 complex were compatible with those of the polynucleosome structure of chromatin. The template activities for transcription were compared in these DNA-histone complexes by separately measuring initiation reaction and chain elongation. The binding of histone H1 to DNA strongly inhibited the initiation, while the binding of the combination of the other four histones to DNA partially inhibited the initiation and chain elongation. The binding characteristics are discussed with regard to the role of histone H1 and the other four histones in chromatin structure and template activity.</p

Absence of C-type virus production in human leukemic B cell, T cell and null cell lines.

Electron microscope observation of cultured human leukemic B cell, T cell and null cell lines and reverse transcriptase assay of the culture supernatants were all negative for the presence of C-type virus. Bat cell line, which propagates primate C-type viruses well, was cocultivated with the human leukemic cell lines, in the hope of amplification of virus if present. Three weeks after mixed culture, the culture supernatants were again examined for reverse transcriptase activity and the cells were tested for syncytia formation by cocultivation with rat XC, human KC and RSb cell lines. All these tests, except for the positive control using a simian sarcoma virus, were negative, suggesting that no C-type was produced from these human leukemic cell lines.</p

Partial purification of Simian virus 40 large T antigen by immunoaffinity chromatography and its detection by enzyme-linked immunosorbent assay.

Simian virus 40 (SV40) large T antigen was partially purified from small amounts of SV40-infected and SV40-transformed cells by immunoaffinity chromatography with high recovery. T antigen, in both crude and partially purified states, was detected rapidly by a sensitive and quantitative enzyme-linked immunosorbent assay (ELISA). Stability of the partially purified T antigen was found to increase by addition of 0.01% bovine serum albumin (BSA).</p