Mutagenic specificity of a novel T4 DNA polymerase mutant.
نویسندگان
چکیده
The in vivo mutational specificity of a novel T4 DNA polymerase mutator mutant, tsM19, was determined. Two genetic tester systems were used to characterize the mutant. Results of our studies indicate that tsM19 promotes transition and transversion mutagenesis and, possibly, frameshift mutagenesis. Central G:C base pairs in runs of three or more consecutive G:C base pairs may be target sites for tsM19-induced transitions.
منابع مشابه
Efficient site directed in vitro mutagenesis using ampicillin selection.
A novel plasmid vector pSELECT-1 is described which can be used for highly efficient site-directed in vitro mutagenesis. The mutagenesis method is based on the use of single-stranded DNA and two primers, one mutagenic primer and a second correction primer which corrects a defect in the ampicillin resistance gene on the vector and reverts the vector to ampicillin resistance. Using T4 DNA polymer...
متن کاملA major role for bacteriophage T4 DNA polymerase in frameshift mutagenesis.
T4 DNA polymerase strongly influences the frequency and specificity of frameshift mutagenesis. Fifteen of 19 temperature-sensitive alleles of the DNA polymerase gene substantially influenced the reversion frequencies of frameshift mutations measured in the T4 rII genes. Most polymerase mutants increased frameshift frequencies, but a few alleles (previously noted as antimutators for base substit...
متن کاملEngineering processive DNA polymerases with maximum benefit at minimum cost
DNA polymerases need to be engineered to achieve optimal performance for biotechnological applications, which often require high fidelity replication when using modified nucleotides and when replicating difficult DNA sequences. These tasks are achieved for the bacteriophage T4 DNA polymerase by replacing leucine with methionine in the highly conserved Motif A sequence (L412M). The costs are min...
متن کاملConstruction and characterization of a bacteriophage T4 DNA polymerase deficient in 3'-->5' exonuclease activity.
Bacteriophage T4 DNA polymerase has a proofreading 3'-->5' exonuclease that plays an important role in maintaining the accuracy of DNA replication. We have constructed a T4 DNA polymerase deficient in this exonuclease by converting Asp-219 to Ala. The exonuclease activity of the mutant T4 DNA polymerase has been reduced by a factor of at least 10(7), but it retains a polymerase activity whose k...
متن کاملA Nested-Splicing by Overlap Extension PCR Improves Specificity of this Standard Method
Background: Splicing by overlap extension (SOE) PCR is used to create mutation in the coding sequence of an enzyme in order to study the role of specific residues in protein’s structure and function. Objectives: We introduced a nested-SOE-PCR (N –SOE-PCR) in order to increase the specificity and generating mutations in a gene by SOE-PCR. Materials and Methods: Genomic DNA from Bacillus thermo...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
عنوان ژورنال:
- Genetics
دوره 106 3 شماره
صفحات -
تاریخ انتشار 1984