Open AccessIn vitroconversion of MVM parvovirus single-stranded DNA to the replicative form by DNA polymerase a from Ehrlich ascites tumour cells
Author(s) -
E A Faust,
Catherine D. Rankin
Publication year - 1982
Publication title -
nucleic acids research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 9.008
H-Index - 537
eISSN - 1362-4954
pISSN - 0305-1048
DOI - 10.1093/nar/10.14.4181
Subject(s) - biology , dna polymerase , aphidicolin , microbiology and biotechnology , dna , polymerase , parvovirus , dna synthesis , dna clamp , dna polymerase ii , base pair , dna replication , in vitro , biochemistry , polymerase chain reaction , virology , virus , reverse transcriptase , gene
A partially purified preparation of DNA polymerase alpha, obtained from the cytosol of Ehrlich ascites tumour cells, has been found to catalyze the conversion of MVM parvovirus, SS DNA (5 kilobases) to RF in vitro. The reaction initiates at a natural 55 base pair hairpin which exists at the 3' terminus of MVM SS DNA. The SS leads to RF conversion is sensitive to aphidicolin, resistant to ddTTP and is promoted by purine ribonucleoside 5' triphosphates, a phenomenon which could not be explained simply by stabilization effects on the in vitro deoxynucleotide precursor pool. In the absence of rNTPs, nascent complementary strands frequently terminate prematurely at a preferred location, between 1300 and 1700 nucleotides from the initiating 3' hairpin terminus. This in vitro system, involving self-primed parvovirus DNA synthesis, provides a convenient assay for those components of the mammalian replicative DNA polymerase complex which are required for the elongation of nascent DNA chains.