Determination of twist and handedness of a 39-base pair segment of DNA in solution

Iwamoto, Satori; Hsu, Ming

doi:10.1038/305070a0

Cited by 7 publications

(2 citation statements)

References 11 publications

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“…The distribution shown in Fig.5 suggests a fundamental unit of supercoiled loop equal to about 12% of Ad5 genome length. The handedness of supercoiling was determined by shadowing the molecules unidirectionally (14) and examined the direction of crossover of supertwisted DNA in comparison with Form I SV40 DNA included in the crosslinked Ad5 virion DNA as an internal standard.…”

Section: Figmentioning

confidence: 99%

Linear adenovirus DNA is organized into supercoiled domains in virus particles

Wong

Hsu

1989

Nucl Acids Res

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Electron microscopic analysis of bis-psoralen crosslinked adenovirus type 5 virion DNA revealed supercoiled domains in an otherwise linear DNA. The existence of supercoiled arrangement in all the virion DNA was demonstrated by the sensitivity of Ad5 DNA in pentonless virus particles to the supercoiling-dependent endonucleolytic activity of Bal31 and S1 nucleases. These nucleases were found to cleave Ad5 virion DNA at specific sites. The observation of stable cleavage sites in the limit digestion of virion DNA by Bal31 suggests that cleavage sites represent boundaries of core proteins which impede the exonuclease activity of Bal31. These data suggest that specific arrangement of core proteins on Ad5 virion DNA. Based on this analysis we determined positions of core proteins in viral genome using indirect end labeling technique. The size of supercoiled domains of virion DNA was estimated by electron microscopy and also by boundaries of mutually exclusive Bal31 cleavage sites at limit digestion condition. Our data suggest each supercoiled domain is equal to about 12% of Ad5 genome length and about 8 loops can be accommodated in Ad5 virion. However sequences at two extreme ends of the viral genome were found to be outside of supercoiled domains. An interesting correlation between supercoiled domains and gene domains of Ad5 genome was noticed.

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Section: Figmentioning

confidence: 99%

Linear adenovirus DNA is organized into supercoiled domains in virus particles

Wong

Hsu

1989

Nucl Acids Res

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“…The single-stranded DNA was checked for possible deletions and rearrangements within the cloned region by hybridization of single-stranded clones containing cloned inserts of opposite orientations (22). The double-stranded regions were then measured by electrophoretic sizing of the Si nuclease (Boehringer Mannhein Biochemicals)-resistant regions or by electron microscopy.…”

Section: Methodsmentioning

confidence: 99%

Transcription unit mapping in adenovirus: regions of termination

Iwamoto¹,

Eggerding²,

Falck-Pederson³

et al. 1986

J Virol

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Using a series of single-stranded clones of adenovirus DNA, we determined the extent of RNA polymerase II transit early in infection for two rightward-reading transcription units. RNA synthesis beginning at the major late promoter (16.5 on the genomic map) continued until approximately 65 to 70 map units so that differential choices of mRNAs within that region were not based primarily on transcriptional decisions but rather on posttranscriptional decisions. Transcription from the major late promoter beginning at 16.5 map units, however, did greatly decrease before approximately 75 map units, ensuring that no mRNAs were formed with sequences beyond approximately 75 map units. Early transcription from E3 then began just past 75 map units (at a higher rate than transcription from the major late promoter); E3 transcripts terminated at least 2 kilobases downstream from a second and final poly(A) site in this transcription unit. The effectiveness of termination in E3 was greater than 95 to 99%.

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