Polyacrylamide gel electrophoresis of influenza virus RNA

Pons, Marcel W.; Hirst, George K.

doi:10.1016/0042-6822(68)90257-2

Cited by 151 publications

(48 citation statements)

References 4 publications

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“…After they were cooled to room temperature, all samples were diluted to 1 ml in STE buffer (9) and divided into equal fractions, one of which was treated with 10 lxg of RNase A and 1 ug of RNase T1 (15 min at 370C). Acid precipitation and counting of radioactivity were as described in Table 1. fortuitously "trapped" in association with the viral ribonucleoprotein at the time of viral maturation.…”

Section: Discussionmentioning

confidence: 99%

“…The method of Pons and Hirst (9) was used with slight modifications. Crude virus pools (250-500 ml), freed of gross cellular debris, were initially clarified by centrifugation at 10,000 X g for 10 min; virus was subsequently pelleted in a Spinco 42 rotor for 30 min at 95,500 X g. All further steps were as described (9). Purified influenza virus stocks in Trisc HCl buffer usually contained 1-2 mg of viral protein per ml [Lowry determination (10) ] and a minimum of 106 hemagglutinating units per ml as measured by standard methods (7).…”

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confidence: 99%

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RNA-Dependent RNA Polymerase Activity Associated with Virions and Subviral Particles of Myxoviruses

Chow

Simpson

1971

Proc. Natl. Acad. Sci. U.S.A.

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In vitro RNA synthesis is catalyzed by an enzyme found in purified virions of each of nine influenza strains tested. The extent of ['HJUTP incorporation observed is strain dependent, with an obligate requirement for Mn'+ ions and RNA template. At least 30% of purified product RNA shows resistance to degradation by ribonuclease A and Ti before and after annealing with exogenous RNA from influenza virions.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

RNA-Dependent RNA Polymerase Activity Associated with Virions and Subviral Particles of Myxoviruses

Chow

Simpson

1971

Proc. Natl. Acad. Sci. U.S.A.

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“…Along with the realization that the influenza A genome was segmented, it was shown that genetic markers in influenza viruses fell into distinct complementation groups (Duesberg, 1968;Pons & Hirst, 1968). It was noted that although the genes of co-infecting viruses readily complemented each other, even when the parent viruses did not have plaque-forming ability in their own right (Barry, 1961;Hirst, 1973;Hirst & Pons, 1973), the progeny virions seldom carried more than one allele at a specific locus (Laver & Downie, 1976;Lubeck et al, 1979;Nakajima & Sugiura, 1977;Varich et al, 2008).…”

Section: Genome Segmentation: a Mixed Blessingmentioning

confidence: 99%

Genome packaging in influenza A virus

Hutchinson

Kirchbach

Gog

et al. 2009

Journal of General Virology

257

279

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“…The tion in sucrose velocity gradients. The profile of sedimentation showed 14 and 20S peaks as well as RNA sedimenting faster than 28S (Fig. 1, B-F).…”

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confidence: 99%

“…6A but not in Fig. 6B virion,13 14 it is reasonable to conclude that the complete viral genome is synthesized. The detection of seven pieces of double-stranded RNA by treatment with RNase or 2 M NaCl fractionation instead of four or sixe5 16 may be because of better resolution of RNA in the acrylamide gel system used in this investigation.…”

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confidence: 99%

NONPERMISSIVE INFECTIONS OF MAMMALIAN CELLS: SYNTHESIS OF INFLUENZA VIRUS GENOME IN HeLa CELLS

Lerner¹,

Hodge²

1969

Proc. Natl. Acad. Sci. U.S.A.

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Abstract.-The RNA synthesis of a single-stranded, multisegmented viral genome has been investigated in nonpermissive host cells. In HeLa cells, both single-and double-stranded influenza viral RNA were made and accumulated, and synthesis was completed by 15 hours. The evidence indicated that all pieces of the genome were synthesized. At least as much viral RNA was synthesized in HeLa cells as in permissive chick embryo fibroblasts cells. Explanations for this nonpermissive infection based on reduced synthesis of viral RNA or rapid destruction of newly synthesized RNA, uncontrolled synthesis of RNA, or partial synthesis of the genome appear to have been ruled out by this investigation. The persistently high amounts of double-stranded RNA, combined with the reported lack of dispersion of nucleocapsid protein from the nuclear region of the HeLa cell are consistent with a failure of assembly of the virion in this host. This nonpermissive infection of HeLa cells with influenza virus may offer a unique system for the accumulation of the intermediates involved in the assembly of a complex RNA viral genome.The nonpermissive infection of HeLa cells with influenza virus appears to be an interesting system for observing the synthesis and assembly of a viral genome composed of single-stranded RNA segments. In HeLa cells virus-specific proteins are synthesized, as can be detected by the techniques of hemadsorption and fluorescent microscopy. Virtually every HeLa cell can be infected and no infectious virus is formed, regardless of the multiplicity of infection." 2 This failure to form infectious virus differs from the von Magnus phenomenon seen in chick embryo fibroblasts because the latter is related to high multiplicities of infection. Furthermore, unlike the permissive infection of chick embryo fibroblasts cells, the viral nucleocapsid protein remains in the nuclear region in the HeLa cell infected with influenza and other myxoviruses.3 6 In this paper some aspects of the synthesis of influenza viral RNA in HeLa cells are described. The

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Polyacrylamide gel electrophoresis of influenza virus RNA

Cited by 151 publications

References 4 publications

RNA-Dependent RNA Polymerase Activity Associated with Virions and Subviral Particles of Myxoviruses

RNA-Dependent RNA Polymerase Activity Associated with Virions and Subviral Particles of Myxoviruses

Genome packaging in influenza A virus

NONPERMISSIVE INFECTIONS OF MAMMALIAN CELLS: SYNTHESIS OF INFLUENZA VIRUS GENOME IN HeLa CELLS

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