In vitro packaging of bacteriophage T7 DNA synthesized in vitro

Journal Article · Sat Jul 01 04:00:00 EDT 1978 · J. Virol.; (United States)

OSTI ID:6724289

Masker, W E; Kuemmerle, N B; Allison, D P

An in vitro DNA packaging system was used to encapsulate T7 DNA that had been synthesized by extracts prepared from gently lysed Escherichia coli infected with bacteriophage T7 carrying amber mutations in gene 3 or in both genes 3 and 6. Isopycnic centrifugation of density-labeled wild-type DNA was employed in an effort to separate product from template; suppressor-free indicator bacteria were used to eliminate contributions from endogenous DNA or contaminating phage. Additional controls indicated that fragmented DNA is packaged in vitro only with very low efficiency and that the frequency of recombination during packaging is too low to affect interpretation of these experiments. T7 DNA replicated by extracts prepared using T7 mutants deficient in both genes 3 and 6 could be packaged in vitro with an efficiency comparable to that found when highly purified virion T7 DNA was used. When T7 deficient in the gene 3 endonuclease but with normal levels of the gene 6 exonuclease was used, fast-sedimenting concatemer-like DNA structures were formed during in vitro DNA synthesis. Electron microscopy revealed many branched and highly complex DNA structures formed during this reaction. This concatemer-like DNA was encapsulated in vitro with an efficiency significantly greater than that found for DNA the length of a single T7 genome.

• Research Organization: Oak Ridge National Lab., TN
• OSTI ID: 6724289
• Journal Information: J. Virol.; (United States), Journal Name: J. Virol.; (United States) Vol. 27:1; ISSN JOVIA
• Country of Publication: United States
• Language: English

Related Subjects

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59 BASIC BIOLOGICAL SCIENCES
BACTERIA
BACTERIOPHAGES
BIOSYNTHESIS
DNA
DNA REPLICATION
ENCAPSULATION
ENZYMES
ESCHERICHIA COLI
GENE MUTATIONS
GENE RECOMBINATION
IN VITRO
MICROORGANISMS
MUTATIONS
NUCLEASES
NUCLEIC ACID REPLICATION
NUCLEIC ACIDS
ORGANIC COMPOUNDS
PARASITES
PHOSPHOTRANSFERASES
SYNTHESIS
TRANSFERASES
VIRUSES