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Updated: Aug 3, 2026

Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli
Published on: March 16, 2011
Abstract:
The mutagenic effect induced by mitomycin C (MC) 20 mg/ml in early, middle and late G1-phase of Crepis capillaris seeds was modified by DNA synthesis inhibitors--caffeine 5 mM, hydroxyurea (HU) 5 x 10(-3) M, 1-beta-D-arabinofuranosylcytosine (araC) 1 x 10(-3) M and aphidicolin (APC) 0.5 mg/ml in the DNA post-synthesis phase (G2). It was shown that under the action of MC in early G1 and the inhibitors in the G2 the total number of aberrations for fixations is lower in all the cases than under the action of MC only. The action of MC in the middle phase results in the differential effect of the inhibitors. HU and APC were shown to potentiate the effect induced by MC and ara-C to reduce it. In both cases, caffeine at the dose 5 mM had small effect on DNA damages induced by MC in early and middle G1. Under the action of caffeine and HU in the G2-phase the crosslinks induced by MC in the late G1 result in aberrations of chromosomal type, illegal for the S-dependent mutagenesis. On the whole, the process of mutation formation turned out to be sensibilized. The specificity of mutagenesis under the action of MC suggests the appearance of mutations of the chromosomal type after the DNA synthesis in G2 phase.
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