Probing transient protein-mediated DNA linkages using nanoconfinement

We present an analytic technique for probing protein-catalyzed transient DNA loops that is based on nanofluidic channels. In these nanochannels, DNA is forced in a linear configuration that makes loops appear as folds whose size can easily be quantified. Using this technique, we study the interaction between T4 DNA ligase and DNA. We find that T4 DNA ligase binding changes the physical characteristics of the DNA polymer, in particular persistence length and effective width. We find that the rate of DNA fold unrolling is significantly reduced when T4 DNA ligase and ATP are applied to bare DNA. Together with evidence of T4 DNA ligase bridging two different segments of DNA based on AFM imaging, we thus conclude that ligase can transiently stabilize folded DNA configurations by coordinating genetically distant DNA stretches.     

DNA计算技术的发展与应用

DNA计算是近年来发展迅速的新兴计算技术,其与信息、数学、物理和纳米领域密切交叉。其信息并行处理速度快,且具有高度分子并行性的优势,得到了广泛的关注。经过近20年的发展,DNA计算在理论研究和实现方面都取得了十分重要的进展。作为未来新兴的计算技术,DNA计算有着巨大的应用潜力。文章对近年来DNA计算中运用的各类技术进行了总结:(1)基于链置换的DNA计算技术;(2)基于核酶的DNA计算技术;(3)基于瓦片的DNA计算技术;(4)基于纳米颗粒的DNA计算技术;(5)基于SiO2的DNA计算技术;(6)细胞内DNA计算技术;(7)其他DNA计算技术,包括表面DNA计算等。在此基础上,对未来DNA计算技术进行了展望并提出建议。     

Changes in antioxidant enzyme levels and DNA damage during aging

Multiple mechanisms underlie the human aging process, but interest continues in the role that free radicals and antioxidants may play. The concentrations of lymphocyte free radical generation (O₂ ⁻& H₂O₂), DNA damage and antioxidant enzyme levels (glutathione Stransferase, superoxide dismutase and catalase) were evaluated in 110 healthy individuals with an age range of 20–80 years. The antioxidant enzyme levels were significantly less in very old age when compared to young. Moreover, the levels of free radical concentration and DNA damage were increased in the same age group with respect to younger group. Cigarette smoking had a positive relation with free radicals and DNA damage, and inverse relation with antioxidants. On the other hand, body mass was found to have positive relation with free radical generation only. The data indicate that depletion of antioxidant enzyme levels would render the older people more susceptible to free radical stress and DNA damage.     

几种苎麻园土壤微生物总DNA提取方法比较

实验设计对比了苎麻园土壤微生物总DNA6种提取方法。方法Ⅰ采用裂解酶、蛋白酶K、SDS与CTAB处理;方法Ⅱ结合利用CTAB、SDS、LDS和BME;方法Ⅲ综合采用玻璃珠、CTAB和SDS法;方法IV结合利用SDS-GITC-PEG;方法V利用尿素、SDS、LDS和BME综合处理;方法VI则利用试剂盒处理土壤。6种方法都可以提取到23kb左右的DNA片段,经过TENP和PVPP方法预处理的土壤样品,能有效去除腐殖酸等污染物,且提取得到的土壤总DNA完整性好、纯度高,不需要纯化就可用于PCR扩增。但不同方法取得的DNA的片段长度、产量和纯度存在明显差异。通过比较,改进并建立了可用于宏基因组构建的高纯度、大片段DNA的方法。     

Protective effect of DNA-mediated immunization with liposome-encapsulated GRA4 against infection of Toxoplasma gondii

The dense granule protein 4 (GRA4) is a granular protein from Toxoplasma gondii, and is a candidate for vaccination against this parasite. In this study, the plasmid pcDNA3.1-GRA4 (pGRA4), encoding for the GRA4 antigen, was incorporated by the dehydration-rehydration method into liposomes composed of 16 mmol/L egg phosphatidylcholine (PC), 8 mmol/L dioleoyl phosphatidylethanolamine (DOPE), and 4 mmol/L 1,2-diodeoyl-3-(trimethylammonium) propane (DOTAP). C57BL/6 mice and BALB/c mice were immunized intramuscularly three times with liposome-encapsulated pGRA4 to determine whether DNA immunization could elicit a protective immune response to T. gondii. Enzyme-linked immunosorbent assay (ELISA) of sera from immunized mice showed that liposome-encapsulated pGRA4 generated high levels of IgG antibodies to GRA4. Production of primary interferon (IFN)-γ and interleukin (IL)-2 in GRA4-stimulated splenocytes from vaccinated mice suggested a modulated Th1-type response. 72.7% of C57BL/6 mice immunized with liposome-encapsulated pGRA4 survived the challenge with 80 tissue cysts of ME49 strain, whereas C57BL/6 mice immunized with pGRA4 had only a survival rate of 54.5%. When immunized BALB/c mice were intraperitoneally challenged with 10³ tachyzoites of the highly virulent RH strain, the survival time of mice immunized with liposome-encapsulated pGRA4 was markedly longer than that of other groups. Our observations show that liposome-encapsulated pGRA4 enhanced the protective effect against infection of T. gondii. Project supported by the Science Foundation of the Health Bureau of Zhejiang Province, China (Nos. 2003QN003 and 2005A001) and the Science Foundation of the Science and Technology Department of Zhejiang Province, China (No. 2006C13022)     

荧光探针法研究三种参类与DNA的结合作用

人参、党参、西洋参三种参类提取液对DNA的保护作用作了初步的研究。利用EB（溴化乙锭）作为荧光探针,测定人参、党参、西洋参三种参类提取液存在下DNA与EB混合液的荧光积分强度,并把不同参类与DNA相互作用的常数定义为D,根据D的大小讨论了三种参类醇提液的浓度大小对DNA保护作用的影响。结果表明,三种参类的醇提液均能与之相互作用,但作用程度不同。D值越大,参类与DNA作用愈强。样品浓度为50mg/ml时,3种参类对DNA保护作用顺序为：人参＆gt;西洋参＆gt;党参。