Construction of a specific amber codon in the simian virus 40 T-antigen gene by site-directed mutagenesis.
AUTOR(ES)
Rawlins, D R
RESUMO
The site-directed bisulfite mutagenesis technique has been used to construct a specific mutation, am404, at nucleotide position 3124 in the simian virus 40 genome. The mutation was contained within a PstI restriction site (map position 0.27) and prevented cleavage by PstI at that position. Nucleotide sequence analysis of the mutagenized region indicated that only a single base pair change had occurred: a guanosine x cytosine leads to adenine x thymine transition. Comparison of the nucleotide sequence of am404 with the known DNA sequence of simian virus 40 indicted that the mutation in am404 resulted in the conversion of a glutamine codon to an amber codon. am404 could not replicate autonomously when transfected into monkey cells (BSC-40) but did replicate when it was cotransfected with the late deletion helper virus dl1007. On the basis of its position in the T-antigen, gene am404 should produce a T-antigen 24% shorter than the wild-type protein.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=353682Documentos Relacionados
- Site-directed mutagenesis of the simian virus 40 large T-antigen gene: replication-defective amino acid substitution mutants that retain the ability to induce morphological transformation.
- Isolation of temperature-sensitive Abelson virus mutants by site-directed mutagenesis.
- Analysis of the diphtheria tox promoter by site-directed mutagenesis.
- Dimerization of simian virus 40 T-antigen hexamers activates T-antigen DNA helicase activity.
- The use of native T7 DNA polymerase for site-directed mutagenesis.