MutS1 is a key protein involved in mismatch repair system for ensuring fidelity of replication and recombination in Deinococcus radiodurans. The zwf gene encodes glucose-6-phosphate dehydrogenase (G6PD) in the pentose phosphate (PP) pathway, which provides adequate metabolites as precursors of DNA repair. In this study, mutS1 and zwf were disrupted by homologous recombination. The zwf mutant (Δzwf) and the zwf/mutS1 double mutant (Δzwf/mutS1) were sensitive to ultraviolet (UV) light, H2O2, and DNA cross-linking agent mitomycin C (MMC), whereas the mutS1 mutant (ΔmutS1) showed resistance to UV light, H2O2 and MMC as the wild-type strain. Inactivation of mutS1 resulted in a 3.3-fold increase in frequency of spontaneous rifampicin-resistant mutagenesis and a 4.9-fold increment in integration efficiency of a donor point–mutation marker during bacterial transformation. Although inactivation of zwf had no obvious effect compared with the wild-type strain, dual disruption of zwf and mutS1 resulted in a 4.7-fold increase in mutation frequency and a 7.4-fold increase in integration efficiency. These results suggest that inactivation of the PP pathway decreases the resistance of D. radiodurans cells to DNA damaging agents and increases mutation frequency and integration efficiency in the mutS1 mutant background.

从放射性污染的土壤样品中分离出1株具有极端耐辐射的微生物BR501,该菌最适生长温度为30℃,不具有氨苄青霉素、氯霉素、四环素、卡那霉素和利福平等抗性.UV、γ辐射生长曲线的结果显示BR501菌株具有极强的辐射抗性.BR501菌株的16S rDNA序列与多株Deinococcus属菌株有很高的同源性,其中,与D.radiodurans菌株的16S rDNA相似性高达99%.结合BR501菌株的表型和Deinococcus 属具有代表性的D.Radiodurans R1菌株的特征,将该菌株归属为Deinococcus属,初步命名为Deinococcus sp.BR501.

Deinococcus sp.BR501,an extremely radioresistant bacterium may contain two nucleotide excision repair pathways: the UV damage endonuclease β(UvsE)-dependent excision repair pathway and the UvrABC-dependent pathway.And the UvsE(coded by dr1819) and UvrABC(Unit A coded by dr1771) are their key enzymes respectively.PCR primers were designed and homologous genes were cloned and disrupted in vitro according to the completely nucleotide sequence of Deinococcus radiodurans R1 genome.Then PCR production was transformed to BR501,and the disrupted mutants(△dr1771,△dr1819 and △dr1771dr1819) were checked and confirmed by homologous recombination.These mutants and the wild type were irradiated by UV light and exposed to the DNA-damaging agents MMC and H_2O_2.The results showed that these pathways were existed in BR501 and only the two pathway losses could result in increased sensitivity to UV and MMC.

In order to construct an effective approach for wild soybean population evaluation and protection, fifty three SSR primers were used to analyze the genetic diversity of 150 wild soybean plants, which were collected from the 10 natural populations at in-situ conserved region in Xinbin county, Liaoning province. One hundred and twenty three alleles were observed at 53 SSR loci, with an average of 2.3 alleles per locus. The similarity matrix correlation analysis showed that the result of 20~25 SSR loci was significantly correlated with that of 53 SSR loci. So, twenty five SSR primers were selected for further analysis. The expected heterozygosity was 0.317, and most of the genetic diversity was found among different groups (66.7%). The genetic diversity was not evenly distributed and there was rare gene flow between groups. The result indicated that, in order to maintain 95% of the diversity, nearly 40 materials should be collected from different groups.

运用易错PCR介导的随机突变技术使高抗草甘膦的菌株的5-烯醇式丙酮酸-3-磷酸莽草酸合酶 (5-Enolpyruvylshikimate-3-phosphate,EPSPs) 基因发生随机突变,转入EPSPs基因缺陷型的大肠杆菌ER2799中,通过互补试验获得对草甘膦抗性明显降低的突变株AZ100.测序结果发现AZ100的EPSPs基因与未突变基因相比,碱基序列发生了两个位点变化,第792位的碱基由鸟嘌呤(G)突变为胸腺嘧啶(T),第1203位的碱基由胞嘧啶(C)突变为鸟嘌呤(G).氨基酸序列也发生了相应的变化,第264位由甘氨酸(Gly)突变为缬氨酸(Val),第401位由丙氨酸(Ala)突变为甘氨酸(Gly).二级结构预测显示此两个突变的氨基酸的位点恰好位于EPSPs与底物结合的关键部位.这两个氨基酸位点的突变,导致EPSPs在有草甘膦存在的情况下活性降低,从而导致菌株的草甘膦抗性降低.将这两个突变的碱基位点与美国的专利保护位点比较,发现这两个突变位点均不在专利保护范围之内.