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Title Chromatin structure of simian virus 40-pBR322 recombinant plasmids in COS-1 cells
Creator/Author Innis, J.W. ; Scott, W.A.
Publication Date1983 Dec 01
OSTI IdentifierOSTI ID: 5943391
Other Number(s)CODEN: MCEBD
Resource TypeJournal Article
Resource RelationMol. Cell. Biol. ; Vol/Issue: 3:12
Research OrgDept. of Biochemistry, Univ. of Miami School of Medicine, Miami, FL 33101
Subject550200 -- Biochemistry ;550400 -- Genetics; CHROMATIN-- MOLECULAR STRUCTURE;NUCLEOPROTEINS;PLASMIDS-- BIOLOGICAL FUNCTIONS;RECOMBINANT DNA-- ENZYMATIC HYDROLYSIS;SIMIAN VIRUS-- RECOMBINANT DNA; CELL CULTURES;DNA REPAIR;DNA SEQUENCING;DNA-ASE;ENDONUCLEASES;HYBRIDIZATION;MONKEYS;STRAND BREAKS
Related SubjectANIMALS;BIOLOGICAL RECOVERY;BIOLOGICAL REPAIR;CELL CONSTITUENTS;CHEMICAL REACTIONS;DECOMPOSITION;DNA;DNA-ASE;ENZYMES;ESTERASES;FUNCTIONS;HYDROLASES;HYDROLYSIS;LYSIS;MAMMALS;MICROORGANISMS;NUCLEIC ACIDS;ORGANIC COMPOUNDS;PARASITES;PHOSPHODIESTERASES;PRIMATES;PROTEINS;RECOVERY;REPAIR;SOLVOLYSIS;STRUCTURAL CHEMICAL ANALYSIS;VERTEBRATES;VIRUSES
Description/Abstract To study the nucleoprotein structure formed by recombinant plasmid DNA in mammalian cells, nuclei were isolated from COS-1 cells after transfection with a recombinant (pJI1) containing pBR322 sequences and a segment of simian virus 40 containing information for a nuclease-sensitive chromatin structure.^ The nuclei were incubated with DNase I.^ DNA fragments which were the size of linear pJI1 DNA were isolated, redigested with restriction enzymes, fractionated by electrophoresis, and detected by hybridization with nick-translated segments prepared from the plasmid DNA.^ Two DNase I-sensitive sites were detected in the simian virus 40 portion of the plasmid at the same sites that were DNase I sensitive in simian virus 40 chromatin prepared late after infection of African green monkey kidney (BSC-1) cells.^ One site extended from the viral origin of replication to approximately nucleotide 40.^ The 21-base pair repeated sequences were relatively DNase I resistant.^ A second site occurred over the single copy of hte 72-base pair segment present in this plasmid.^These results indicate that the nuclease-sensitive chromatin structure does not depend on the presence of viral structural proteins.^ In addition, late viral proteins added to PJI1-transfected COS-1 cells by superinfection with simian virus 40 caused no change in the distribution of DNase I-sensitive sites in plasmid chromatin.^Analysis of transfected plasmid DNA may provide a general method applicable to the study of the chromatin structure of cloned segments of DNA.
Country of PublicationUnited States
LanguageEnglish
FormatPages: 2203-2210
System Entry Date2001 May 13

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