Mutant profiles of selectable genetic elements.
AUTOR(ES)
Wurst, H
RESUMO
A method is presented that allows simultaneous analysis of the effects of all possible point mutations within a specific mutation window of at least 50 base pairs on a DNA fragment that codes for a selectable function. It relies on the detection of mismatched base pairs with hydroxylamine and osmium tetroxide. A mutant plasmid library of randomly distributed point mutations within the lacZ' gene of Escherichia coli was selected for functional alpha-complementation by growth on lactose. The DNA fragments of the selected and unselected library were each heat denatured and again renatured, thereby generating a randomly distributed set of all possible mismatches within the mutagenesis window. Cytidine-containing mismatches were then detected with hydroxylamine, and thymidine-containing mismatches were detected with osmium tetroxide. When this procedure was performed for both DNA strands, all mismatches could be detected. A comparison of the results of the unselected and selected library leads to an estimation of the effects of each detectable mutation on alpha-complementation in vivo. This method, called "mutant profiling," should be applicable to all selectable genetic elements.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=52836Documentos Relacionados
- Genetic definition of ras effector elements.
- Inhibition of bacteriophage replication by extrachromosomal genetic elements.
- Mammalian cell function mediating recombination of genetic elements.
- Structural similarities between viroids and transposable genetic elements.
- Polyomavirus origin for DNA replication comprises multiple genetic elements.