Studies On The Mutation Mechanism Stimulated By Low Energy N～+ion Beam Implantation: Analyzing Mutational Spectrum

Base on the summarizing of studies on mutagenic mechanisms, gene transfering, varieties industrial microbes breeding with low energy ion beam, a salmonella typhimurium strain carrying a super-repressed mutant of purR(purR^S) was used as an experimental system for study mutation mechanisms caused by low energy N⁺ ion beam implantation.PurR encodes a repressor protein controlling expression of purine biosynthetic genes. Because the expression of lac genes fused with purD was highly repressed by super-repressor of purR. This kind of purR^S mutants can't grow on the NCE plate with lactose as sole carbon source. But if regulatory gene purR ox purD operator's cis-acting element PUR box produced mutatins, the cells can free from super-repressing state, which let it to grow on the NCE-lacose medium. Screen out mutants by ion beam implanting and andanalyze mutational spectrum.Dry logarithmic purR^S were implanted by the N⁺ ion beam of 15keV, 110(2.6×10¹³ions/cm²) dose, under vacuum state. Via genetic identify the 100 mutants weharvested, 80purR^- and 20 PURbox mutants were included. Clone target segment of PURboxmutants to analyse the nucleotide sequences and mutational spectrum. We report the followresults and conclutions.(1)Bacteria in aseptic water were dried up in 35' in laminar flow cabinet. The survival rate was 9.7%. 0.85% survived after the whole experiment time, 55'. Survival rate under vacuum is more than 50% and implantation survival rate is 20.6%⁴5.2% with 110 dose. Mutagenesis rate are higher than that of spontaneous mutation by 7.0⁹.3 times.(2)Proportion of purR and PURbox mutants in mutagenesis is 0.8:0.2.(3)The spectra anlysis of mutagenized mutants of PURbox showed that all the mutation events were base substitution, occurred at three conserved site: C8→A G9→T, C14→T. Proporty of transition and transvertion in mutagenized mutaion is 0.15:0.85. There is a hotspot: G9→T. 75 percent mutants belonged. Mutagenesis mutational spectra is narrower than other two kinds of mutations: grow-depended and adaptive mutation. The mutagen mechanism is presumed single factor dominant.(4)There was 85 percent transvertion in mutagen spectra, and they were all G:C→T:A transvertion. The maximal possibility of mutagen mechanism is analyzed as followed:ion beam implant → breakage cell surface and enter the interior → stimulate plentiful radical born → radical oxidize and damnify DNA → G becomes 8—oxoG → 8—oxoG pairing A → result in G:C→T:A transvertion(5)Production of transition was presumed that ion beam implantion damnified DNA, stimulate activity of MMR interveniented by methyl reducing. Mutation event stimulated by MMR activity reducing is mainly transition, transvertion is very infrequently. DNA damnification or replication stagnation brings on ssDNA, which as signal activate SOS reaction. It relieves dinB( encodes Polâ…£). Polâ…£ expression induces a mass of errors which gobeyond repaired.