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Biomedical subjects

S Mak

Publications and source records attributed to S Mak.

At least 55 records · Page 3Linked to original sources
PubMed1976

Purification of adenovirus messenger ribonucleic acid by an aqueous polymer two-phase system.

An aqueous polymer phase system containing 6.3% (w/w) dextran and 3.5% (w/w) poly(ethylene glycol) in 10 mM phosphate buffer (pH 8.0) was developed to select RNA-DNA hybrids from unhybridized RNA. The top phase of this phase system, which contains DNA and the RNA-DNA hybrids, can be used to purify adenovirus messenger RNA both early and late in the infectious cycle. The hybrids can be melted by heat in the top phase and the messenger RNA selected by oligo(dT)cellulose chromatography whereupon the polymers and the DNA percolate and the polyadenylated messenger RNA absorb to the column. The isolated messenger RNA appears to be almost quantitatively recovered at a purity from 70 to 90% depending on the concentration of the specific messenger RNA in the starting material. Early and late viral messenger RNA were selected on the complementary strands of adenovirus DNA according to this procedure.

Adenoviruses, Human↗
PubMed1976

Adenovirus type 12-specific RNA sequences during productive infection of KB cells.

The complementary strands of adenovirus type 12 DNA were separated, and virus-specific RNA was analyzed by saturation hybridization in solution. Late during infection whole cell RNA hybridized to 75% of the light (1) strand and 15% of the heavy (H) strand, whereas cytoplasmic RNA hybridized to 65% of the 1 strand and 15% of the h strand. Late nuclear RNA hybridized to about 90% of the 1 strand and at least 36% of the h strand. Double-stranded RNA was isolated from infected cells late after infection, which annealed to greater than 30% of each of the two complementary DNA strands. Early whole cell RNA hybridized to 45 to 50% of the 1 strand and 15% of the h strand, whereas early cytoplasmic RNA hybridized to about 15% of each of the complementary strands. All early cytoplasmic sequences were present in the cytoplasm at late times.

Adenoviridae↗
PubMed1974

Comparative studies on functions of human adenovirus type 12 and its low oncogenic mutant virions.

Physical and biological properties of highly oncogenic human adenovirus type 12 were compared with a low oncogenic mutant (cyt mutant). Parental and cyt mutant virions had very similar density and DNA size. However, the parental strain virion preparations contained a much higher proportion of defective virions (capable of cell killing, but not able to induce T- or V-antigen) than cyt mutant stock. It was also found that cyt mutant had a reduced virus yield in several human cell lines compared with the parental strain.

Adenoviridae↗
PubMed1971

Defective virions in human adenovirus type 12.

Purified preparations of human adenovirus type 12 showed two bands when subjected to isopycnic centrifugation in a density gradient of cesium chloride. Their density difference was about 0.003 g/ml, suggesting a small difference in their deoxyribonucleic acid to protein ratio. Virions with a lighter density can kill human KB cells and induce T antigen as efficiently as the heavy virions. However, they appeared incapable to form plaques. Two passages of the heavy infectious virions at low multiplicity of infection did not produce significant amounts of light virions; however, when it was passed at high multiplicity of infection, the light band became visible in a cesium chloride density gradient.

Adenoviridae↗
PubMed1970

Functional heterogeneity of virions in human adenovirus types 2 and 12.

Purified preparations of adenovirus types 2 and 12 were used to infect KB cells at different input multiplicities. The resulting infected cultures were scored for inclusion body formation, production of infectious centers, and cloning efficiency. Both preparations were found to contain some defective particles capable of preventing a cell from cloning but unable to induce inclusion bodies or form plaques. The proportion of such defective particles in adenovirus 12 was about 10 times that in adenovirus 2. At high input multiplicities, the percentage of cells displaying an inclusion body was less than that predicted by the Poisson distribution and reached a maximum of 40 to 60% for adenovirus 2 and 12 to 15% for adenovirus 12. This reduction may be due to interference by large numbers of non-plaque-producing particles infecting each cell. The per cent of cells forming infectious centers was substantially greater for adenovirus 2 than for adenovirus 12 when compared at the same input plaque-forming units, reaching a maximum of 35 to 73% for adenovirus 2 and 5 to 10% for adenovirus 12. The low value for adenovirus 12 may be a result of the same interference phenomenon.

Adenoviridae↗
PubMed1969

Transcription and replication of viral deoxyribonucleic acid in cells coinfected with adenovirus types 2 and 12.

The yield of infectious virus was determined for KB cells infected with both adenovirus types 2 (ad 2) and 12 (ad 12). It was found that the yield of the former was greatly reduced, whereas that of the latter was not affected significantly. The reduction in virus yield was accompanied by an inhibition of ad 2 virus-specific ribonucleic acid (RNA) and viral deoxyribonucleic acid (DNA) synthesis at various times after infection. On the other hand, the rate of synthesis of ad 12 virus-specific RNA and viral DNA was not inhibited, but advanced in time. The total amount of ad 12 viral DNA synthesized was not affected by coinfection with ad 2. These results suggest that ad 2 infection hastens the maturation of ad 12.

Adenoviridae↗
PubMed1968

Biochemical studies on adenovirus multiplication. 13. Synthesis of virus-specific ribonucleic acid during infection with human adenovirus type 12.

The transcription of virus-specific ribonucleic acid (RNA) was studied in KB cells infected with adenovirus type 12 (strain Huie). Viral deoxyribonucleic acid (DNA) synthesis began at 12 to 15 hr after infection, and virus maturation occurred between 20 and 50 hr after infection. The rate of incorporation of (3)H-uridine into RNA per infected cell was stimulated, reaching a maximum of 1.6 times that of uninfected cells at 20 hr after infection. "Early" viral messenger ribonucleic acid (mRNA) constituted 0.1% of RNA synthesized at 11 hr, and "late" viral mRNA constituted 50% of RNA synthesized at 45 hr after infection, as determined by hybridization of viral DNA with labeled RNA from infected cells pulse labeled with (3)H-uridine. The species of virus-specific RNA synthesized at 22 hr after infection (when virus maturation has just begun) and at 45 hr (when virus maturation is nearly complete) were studied further: (i) 22- and 45-hr RNA had the same average guanine plus cytosine content, 47%, (ii) 22- and 45-hr RNA contained mostly the same viral nucleotide sequences, (iii) 45-hr RNA had a five times higher concentration of virus-specific RNA molecules than did 22-hr RNA, and (iv) 22- and 45-hr RNA contained virus-specific nucleotide sequences transcribed from all, or nearly all, of the viral genome.

Adenoviridae↗