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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)
 
 
 
 
 

Multiple point mutations in a shuttle vector propagated in human cells: evidence for an error-prone DNA polymerase activity.

Mutagenesis was studied at the DNA-sequence level in human fibroblast and lymphoid cells by use of a shuttle vector plasmid, pZ189, containing a suppressor tRNA marker gene. In a series of experiments, 62 plasmids were recovered that had two to six base substitutions in the 160-base-pair marker gene. Approximately 20-30% of the mutant plasmids that were recovered after passing ultraviolet-treated pZ189 through a repair-proficient human fibroblast line contained these multiple mutations. In contrast, passage of ultraviolet-treated pZ189 through an excision-repair-deficient (xeroderma pigmentosum) line yielded only 2% multiple base substitution mutants. Introducing a single-strand nick in otherwise unmodified pZ189 adjacent to the marker, followed by passage through the xeroderma pigmentosum cells, resulted in about 66% multiple base substitution mutants. The multiple mutations were found in a 160-base-pair region containing the marker gene but were rarely found in an adjacent 170-base-pair region. Passing ultraviolet-treated or nicked pZ189 through a repair-proficient human B-cell line also yielded multiple base substitution mutations in 20-33% of the mutant plasmids. An explanation for these multiple mutations is that they were generated by an error-prone polymerase while filling gaps. These mutations share many of the properties displayed by mutations in the immunoglobulin hypervariable regions.[1]

References

  1. Multiple point mutations in a shuttle vector propagated in human cells: evidence for an error-prone DNA polymerase activity. Seidman, M.M., Bredberg, A., Seetharam, S., Kraemer, K.H. Proc. Natl. Acad. Sci. U.S.A. (1987) [Pubmed]
 
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