极端耐辐射奇异球菌Deinococcus sp.BR501切除修复系统突变株的构建及功能的鉴定

极端辐射异常球菌Deinococcussp.BR501的核苷酸切除修复系统,可能包含两条途径:依赖于UV损伤内切酶β(UvsE)的修复途径和依赖于UV损伤内切酶α(UvrABC)的修复途径,其关键酶分别为UvsE(dr1819编码)和UvrABC(A亚基dr1771编码)。根据Deinococcus radiodurans的序列,采用同源克隆的方法,设计PCR引物、克隆基因和体外阻断目的基因,再通过PCR产物线性转化的方法,筛选到同源双交换的重组阻断突变株△dr1771、△dr1819和△dr1771与△dr1819的双突变株。对此突变株和野生型均进行UV辐射抗性和化学损伤试剂MMC与H2O2的损伤修复能力的分析。结果表明了BR501中两条核苷酸切除修复途径的存在,并且只有两条途径同时缺失时,才能使菌体对UV和DNA交联剂MMC敏感。     

耐辐射球菌中priA类似基因突变对DNA修复和DNA转化的影响

DNA损伤很易阻断复制叉的前进。损伤DNA的修复以及接下来停止复制叉的重启动过程对细胞生存极为重要。依赖于PriA的复制重启动机制是细菌复制重启动的主要途径。为了解priA类似基因Dr2606在耐辐射球菌中的作用,并检测Dr2606在DNA修复中的作用,本研究用卡那霉素抗性基因代替Dr2606阅读框,构建了Dr2606缺失突变株,并对突变株进行UV和丝裂霉素处理,测定了Dr2606突变株的转化效率。Dr2606的突变导致菌体生长缓慢,细胞生存率急剧下降。意外的是,耐辐射球菌的DNA修复能力没有削弱。但突变株的转化效率大大削弱。这说明在耐辐射球菌中priA类似基因Dr2606对停止复制叉的重启动过程并不是必需的;耐辐射球菌不依赖于原点的复制重启动过程可能与其他细菌不同。     

不同利福平浓度压力下耐辐射球菌的自发突变率与突变谱研究

根据耐辐射球菌(Deinococcus radiodurans) rpoB基因的保守性,建立了rpoB/Rifr 突变分析系统。为研究不同利福平(rifampin, Rif)浓度对耐辐射球菌自发突变率和突变谱的影响,分别检测耐不同利福平浓度下辐射球菌的突变率。结果表明：在5μg/ml的利福平浓度下耐辐射球菌的突变率显著高于其在25和50μg/ml浓度下的突变率。随利福平浓度的不同而表现出的突变谱差异,表明在不同利福平浓度引起的活性自由氧(reactive oxygen species, ROS)压力下,耐辐射球菌可能使用不同抗突变的应对策略。在低利福平浓度下,耐辐射球菌重点修复碱基替换突变导致的DNA损伤。这可能是因为ROS的主要作用是导致DNA出现氧化性碱基损伤。     

低剂量超辐射敏感与诱导辐射抗性的研究进展

低剂量超辐射敏感(HRS)和诱导辐射抗性(IRR)是辐射生物学的两个重要现象,研究这两个现象发生的机理对于辐射保护、控制肿瘤发生、研制新的功能材料具有重要的意义。辐射产生的DNA双链断裂(DNA double-strand breaks,DSBs)是辐射诱导细胞失活的主要损伤形式。对DSBs不能进行修复或者不能进行正确的重新连接是引起细胞死亡的主要原因。与DNA合成前期(first gap phase,G1期)和DNA合成期(synthetic phase,S期)相比,HRS在DNA合成后期(second gap phase,G2期)的表现非常明显。HRS和IRR之间相互转换调控的机制可能是G2期细胞中的一部分存在一个激活过程的发生。DNA损伤修复机制在HRS/IRR过程中发挥了重要作用。在低于20cGy的剂量条件下,细胞产生效率较低的损伤效应,细胞死亡迅速增加,低剂量超辐射敏感现象(HRS)出现;在高于20cGy的条件下,辐射造成DNA双链断裂,这些DSB本身或者由辐射引起的DNA变化激活了ATM(ataxia telangiectasia mutated),G2期专有的检测点被激活,细胞周期在G2期停顿下来,激活的G2期专有的检测点促进对DNA进行修复,提高了细胞的存活率,此时,诱导辐射抗性(IRR)出现。尽管这些结果使人们对HRS/IRR有了一个比较清晰地了解,目前仍然存在着一些关键问题有待解决。例如,为什么有些细胞系表现HRS/IRR现象,而有些细胞系不表现HRS/IRR现象;细胞中是否存在针对单一剂量的突变的HRS/IRR等。这些问题可能是将来重要的研究方向。     

耐辐射球菌清除活性氧自由基及对DNA的保护作用

用化学发光法分析了耐辐射球菌 (D .radiodurans)的两个菌株 (R1、KD830 1 )和大肠杆菌对超氧阴离子自由基、羟基自由基和过氧化氢的清除能力。通过羟基自由基诱导的DNA氧化损伤化学发光模型考察了耐辐射球菌细胞提取物对自由基氧化导致DNA氧化损伤的保护作用。结果表明 ,耐辐射球菌提取物能显著清除O-2·、H2 O2 和·OH ,清除能力明显高于大肠杆菌 ,并能有效地防止自由基所致DNA氧化损伤 ,表现出超强的抗氧化和耐辐射能力 ,这归因于耐辐射球菌中具有较高的抗氧化酶活性     

耐辐射球菌Rsr增强大肠杆菌抗逆性的研究

耐辐射球菌(Deinococcus radiodurans)Rsr蛋白(DR_1262编码)是一种对辐射抗性起重要作用的RNA结合蛋白.本研究将完整的rsr基因连接到带有groESL启动子的pMD-GroE载体上,构建了Rsr表达菌株.以合空载质粒pMD-GroE的大肠杆菌为对照,发现Rsr的表达提高了大肠杆菌的UV辐...     

奇球菌属的最新研究进展及其应用

奇球菌属(Deinococcus)是迄今为止发现的最具抗辐射能力的一类细菌,在环境治理上具有巨大的应用前景,其中的耐辐射奇球菌(Deinococcus radiodurans)还是研究DNA损伤修复和极端抗性应激机制的模式生物。本文介绍了奇球菌属最新的研究进展,对奇球菌属生理结构特点和抗性机制进行了讨论,并对其在生物技术上的应用前景进行了展望,以期为后续研究者们快速了解该属细菌及其在生产上的应用提供支撑。     

杨树葡萄糖-6-磷酸脱氢酶（G6PDH）基因启动子的克隆与分析

葡萄糖-6-磷酸脱氢酶是磷酸戊糖途径的关键性调控限速酶,其主要功能是为脂肪酸合成、氮还原和谷胱甘肽等生物分子合成提供还原力NADPH,也为核酸合成提供戊糖;此外,还参加非生物逆境胁迫应答反应。因此,G6PDH对植物的生长发育起着非常重要的作用。本文利用甜杨G6PDH基因和毛果杨基因组序列,通过PCR获得了甜杨G6PDH基因上游1400bp的序列。序列分析结果表明,该序列具有启动子的基本元件TATA-box、CAAT-box。此外,还包含多个胁迫诱导元件,如低温诱导元件LTR,盐诱导元件GT-1,抗冻、缺水、脱落酸、抗寒元件MYB和MYC,以及光响应元件L-box、G-box、3AF-1、TC丰富区等。     

同步辐射(软X射线)对冬小麦的诱变效应及机理研究Ⅲ.辐照种子产生的自由基和辐照引起种胚DNA非按期合成的研究

应用电子自旋 (ESR)波谱术研究了同步辐射辐照种子产生的自由基及其与辐射损伤之间的关系。结果表明 ,同步辐射辐照的种子产生了大量的自由基 ,其种类、性质不同于γ射线 ,短寿命自由基 (小于 48h)产额高于γ射线 ,长寿命自由基 (大于48h)的稳定性大于γ射线。同步辐射自由基产额与辐照剂量呈线性关系。用3H TdR掺入法研究了受辐照种胚萌发早期DNA的合成动态 ,结果表明 ,同步辐射能够引起种胚DNA的损伤修复合成 ,其特点不同于γ射线。原冬 6号和北京41 1品种的DNA修复合成出现在种胚吸水后第一个正常的DNA合成峰之前 ,京冬8号品种出现在第一个小峰之后和 2个高峰之前     

枯草芽孢杆菌sacB基因的功能验证及应用

本研究将枯草芽孢杆菌(Bacillus subtilis)基因组DNA中克隆的1.4 kbsacB基因片段连接到表达载体pET-28a(+),构建了sacB基因表达载体pSacB。通过测定蔗糖水解产生的葡萄糖具有的还原性,确定sacB基因产物具有蔗糖果聚糖酶活性。该基因表达后,导致大肠杆菌(Escherichia coli)细胞不能在含5%蔗糖的培养基中生存。同时发现sacB基因也能赋予耐辐射菌株Deinococcussp.BR501蔗糖敏感性。利用条件致死sacB基因的这一特性,我们分析了D.sp.BR501的突变率,结果表明DNA错配修复缺失突变株(ΔmutS1)的突变频率明显高于野生型。因此枯草芽孢杆菌sacB基因可作为验证DNA损伤修复能力的一种筛选标记。     

Soil-profile distribution of primary and secondary plant-available nutrients under conventional and no tillage

Nutrient distributions under no tillage (NT) compared with conventional disk-and-bed tillage (CT) management in the warm, humid region of the southeastern USA need to be assessed so that future placement, quantity, and type of fertilizers can be altered, if necessary, to efficiently match crop demands. We determined soil-profile distributions of pH, N, P, S, K, Ca, Mg, Na, Zn, Fe, Mn, and Cu to a depth of 0.9 m at the end of 8.5 years of continuous CT and NT management on a Weswood silty clay loam (fine, mixed, thermic Fluventic Ustochrept) in southcentral Texas. Most dramatic changes occurred within the 0–0.05 m depth, where soil under NT had lower pH, Fe, and Cu than under CT, but greater P, K, Zn, and Mn. Greater P and K under NT than under CT also occurred below the till-zone (0.15–0.3 m). At a depth of 0–0.3 m, soil under NT contained greater amounts of extractable P, K, Zn, Fe, Mn, and Cu than under CT. Nitrogen fertilization had little effect on nutrient distributions, except resulting in greater extractable K at 0–0.05 m and greater nitrate at 0–0.15 m. Few changes in soil-profile distributions were observed for extractable S, Ca, Mg, and Na. Long-term continuous use of NT on this fine-textured, high-fertility (except for N) soil had no apparent adverse effects on nutrient distributions relative to CT, but enhanced conservation and availability of P, K, Zn, Fe, Mn, and Cu near the soil surface where crop roots proliferate.     

Assessing the effectiveness of conservation measures: Resolving the “wicked” problem of the Steller sea lion

“Wicked” problems are those that are complex and that change when solutions are applied. Many conflicts in conservation fall into this category. The study approached the problem of how to constrain the apparent wickedness of a problem in the conservation management of a species by using simple empirical indicators to carry out iterative assessment of the risk to a population and to document how this risk evolves in relation to the addition of new data and the implementation of management actions. Effects of high levels of uncertainty within data and also concerning population structure were examined through stochastic simulation and by exploration of scenarios. Historical trends in the example used, the Steller sea lion, showed rapid declines in abundance in some regions during the 1980s. The current total population is 130,000-150,000 Steller sea lions through Alaska and British Columbia and this number has been stable since about 1990 in spite of regional differences in population dynamics. Regional differences in the sequence of changes in the number of pups and non-pups, suggested that an internal re-distribution of juveniles could have happened between 1980 and 1990. Current productivity also appears close to the long-term mean. Stochastic population projection using various scenarios showed that, based upon this history, the risk of extinction for the population has declined and is below reasonable thresholds for considering the population to be endangered. The trends in risk suggest that management actions taken since 1990 have probably been effective. Consequently, the conservation management objectives for the Steller sea lion are probably being met. The approach provides a mechanism, based upon experience and scenario analysis, for exploring future policy options and may help to constrain the debate amongst stakeholders about the cost-benefit trade-offs associated with different options.     

Enhanced establishment and growth of giant cardon cactus in an eroded field in the Sonoran Desert using native legume trees as nurse plants aided by plant growth-promoting microorganisms and compost

To evaluate the feasibility of long-term desert reforestation technology of mixed vegetation, cardon cactus (Pachycereus pringlei) seedlings from indoor and outdoor nurseries were planted in the field adjacent to one seedling of potential legume nurse trees: mesquite amargo (Prosopis articulata), yellow palo verde (Parkinsonia microphylla), and blue palo verde (Parkinsonia florida). Some of the planting holes were also supplemented with common dairy compost. Additionally, the combinations of legume tree–cactus were inoculated with either a consortium of desert arbuscular mycorrhizal (AM) fungi, plant growth promoting bacteria (PGPB; the diazotroph Azospirillum brasilense Cd, and the phosphate solubilizer Paenibacillus sp.), or a mixture of all. The field experiments were evaluated periodically during 30 months for survival and growth. Cardons reared in an outdoor screen house survived better in the field than those reared in a controlled growth chamber and hardened later outdoors. Association with any legume nurse tree increased survival and enhanced growth of untreated cardons. For cardons growing alone, application of either compost, AM fungi, and all the treatments combined increased survival. For these plants, no treatment affected plant growth during the first 3 months after transplanting. Later, all treatments, except for AM fungi, enhanced plant growth. However, only 2 years after transplanting the enhanced growth effect of AM fungi was also significant. In the presence of the legume nurse trees, transient positive effects on cardon growth were recorded. General evaluation after 30 months of cultivation showed that the treatments positively affected cardon growth when growing alone or in combination only with mesquite amargo but not with the other two legume trees. This study proposes that young legume trees have the capacity to enhance survival and growth of cardon cactus, depending on the legume cactus combination. Additional treatments such as compost or PGPB can either amplify the effect or else attenuate it.     

Karyotypic and genome size variation in Cajanus cajan (L.) Millsp. (pigeonpea) and some wild relatives

Detailed karyotypes and 4C DNA amounts have been studied in five cultivars of Cajanus cajan and 20 species belonging to Cajanus, Rhynchosia, Dunbaria, Flemingia and Paracalyx. C. cajan shows intraspecific variability and its karyotype is most similar to that of C. cajanifolius (sect. Cajanus) in the morphology and number of satellite chromosomes and the lack of any correlation between chromosome size within the complement and asymmetry. Karyotypes of C. lineatus and C. sericeus belonging to sect. Atylia are similar with respect to maximum r-index and the ratio of longest and shortest chromosomes in their respective complements. C. acutifolius (sect. Frutocosa) is distinct in having a very low ratio (1.44) between the longest and the shortest chromosomes, while in C. albicans, C. goensis, C. scarabaeoides (sect. Cantharospermum) there is no chromosome pair with r-index >2.0. C. mollis and C. volubilis (sect. Volubilis) show similarity with regard to ratio between the longest and the shortest chromosomes and a distinct chromosome pair with an arm ratio of 2.7 and C. platycarpus is distinct (sect. Rhynchosoides) in having the smallest ratio (1.36) between the longest and the shortest chromosomes. 4C DNA amounts in Cajanus and Rhynchosia vary between 3.28 pg to 11.69 pg and 4.92 pg to 11.13 pg respectively, while in Dunbaria, Flemingia and Paracalyx these vary between 4.70 and 7:06 pg. In the genus Cajanus karyotypic features and 4C DNA amounts agree with the sectional classification and bring out a close relationship between C. cajan and C. cajanifolius. This is supported by studies based on seed protein patterns, isozyme analysis, trypsin and chymotrypsin inhibitor patterns, RFLP and RAPD data and crossability relationships. A clear relation between DNA amount and abnormalities effecting the extent of pairing and recombination in interspecific hybrids shows the importance of this study in developing future breeding programme involving C. cajan and its wild relatives that are of potential value.     

Effect of some fungicide seed treatments on the survival of Bradyrhizobium japonicum and on the nodulation and yield of soybean [Glycine max. (L) Merr.]

Several commercial fungicide seed treatments were evaluated for their possible effect on the survival of Bradyrhizobium japonicum on seeds and on the nodulation and yield of soybeans in a greenhouse and a field experiment. quinolate Pro (carbendazim and oxine copper), Vitavax 200FF (carboxin and thiram), and Monceren (pencycuron) had a small effect or no effect on the survival of B. japonicum and on the nodulation and yield of soybeans. They can thus be considered compatible with soybean seed inoculation. Germipro UFB (carbendazim and iprodione), Apron 35J (metalaxyl), and Tachigaren (hymexazol) decreased B. japonicum survival and the nodulation and yield of soybeans and thus cannot be considered compatible with soybean seed inoculation.     

Einfluss der düngungsmethoden auf die entwicklung von zuckerrübenkrankheiten

Während neunjähriger (1994-2002) Untersuchungen in der Versuchsanstalt Ba?cyny (Fahlerde, Fall-Gleyboden, ausgebildet aus leichtem Lehmboden) wurde der Befall mit Krankheiten an der Zuckerrübe in Abhängigkeit von der Düngungsmethode bewertet. Folgende Methoden wurden angewendet: A - Mineraldünger, B - Stalldung und Mineraldünger, C - Stroh-, Gründung und Mineraldünger, D - Kompostdünger und Biopräparate (ökologische Düngung). Auf den Objekten A, B, C wurden bei konstanter Phosphor-Kalium-Düngung die Stickstoffgaben von 0 bis 180 kg.ha^?1 differenziert. Auf dem ökologischen Objekt D ohne Mineraldünger wurden statt Stickstoff die Biopräparate P₅₀₀ und P₅₀₁ verabreicht. Der geringste Befall von Krankheiten wurde an der Zuckerrübe auf jenem Objekt festgestellt, das mit Kompost und den Biopräparaten gedüngt worden war. Im Rahmen der konventionellen Düngungsmethoden wurde allgemein die höchste Intensität der Krankheiten an der mit Stroh, Gründung und Mineraldünger gedüngten Zuckerrübe beobachtet. Ausnahmen von dieser Regel bildete das Vorkommen von echtem Mehltau und Keimlingsfäule. Auf den Objekten A, B, C nahm die Intensität der Krankheiten mit der Steigerung der Stickstoffgabe zu. Auch die auf Objekt D verabreichten Biopräparate stimulierten die Entwicklung der Krankheiten. The effect of various fertilization methods on the diseases of sugar beet was estimated on the basis of nine-year studies (1994-2002) conducted in the Experimental Production Farm in Ba?cyny on loess-based, sedimentary-gley, light-clay soil. The following fertilization methods were applied: A - mineral fertilization, B - mineral and manure fertilization, C - straw, green manure and mineral fertilization, D - ecological compost and biodynamic preparations fertilization. In the objects A, B and C fertilized with phosphorus and potassium, various doses of nitrogen ranging from 0 to 180 kg/ha^?1 were used. In the ecological object D, where mineral fertilization was not used, biodynamic preparations P₅₀₀ and P₅₀₁ were applied instead of nitrogen. The lowest ratio of the diseases of sugar beet was observed in the object fertilized with the compost and biodynamic preparations. In view of the conventional fertilization methods, generally the highest incidence of diseases was noted in the sugar beet fertilized with straw, green manure and mineral fertilizers. The exception was the occurrence of powdery mildew and the black leg. In objects A, B and C the incidence of the diseases increased with the increase in nitrogen doses. The applied biodynamical preparations in the object D stimulated the development of diseases too.     

Prospects for control of Pythium damping-off of lettuce with Trichoderma,Gliocladium and Enterobacter spp.

Summary Damping-off of lettuce (Lactuca sativa) caused by Pythium ultimum was studied in pots containing a non-sterile potting mix in the glasshouse. Fifty P. ultimum sporangia g^-1 compost reduced the plant stand to 15% and shoot dry weight to 18%, but this reduction was totally prevented by applying Trichoderma harzianum at 2×10⁵ viable propagules g^-1 potting mix. Gliocladium virens and Enterobacter cloacae also alleviated the damping-off. E. cloacae did not adversely affect the action of the fungal antagonists. The co-existence of the bacterial and fungal antagonists was revealed on the root surface and inner surface of the testa by scanning electron microscopy, indicating the compatibility of the biocontrol agents.     

细菌、真菌及植物氮营养信号研究进展

氮素吸收代谢是所有生物生命活动的核心部分,因此,各种生物对氮的吸收与同化过程都有精细的调节。这种调节依赖于生物对体内氮素状况信号及体外（生长介质中）氮素信号的感受。过去几十年中,在细菌、真菌中已经对氮素营养信号有了较好的研究,而植物中则相对缓慢,但也有了一定的认识。本文重点比较细菌、真菌及植物中氮信号系统的组成,以期为进一步认识植物中的氮信号接受与转导系统提供启示。     

Aircraft operations near concentrations of birds in Antarctica: The development of practical guidelines

Aircraft operations have the potential to disturb and to impact negatively on bird life. A gradient of increasing behavioural response is evident in birds when exposed to increasing aircraft stimulus. The most major disturbance is likely to lead to impacts on the health, breeding performance and survival of individual birds, and perhaps bird colonies. A process of revision to policies on aircraft operations contained in management plans for a number of specially protected areas in Antarctica by the United Kingdom, accompanied by consultations made within the scientific community through the Scientific Committee on Antarctic Research (SCAR) and with operational interests through the Council of Managers of National Antarctic Programmes (COMNAP) resulted in new guidelines being adopted by the Antarctic Treaty Consultative Parties in June 2004. The principal recommendations of the guidelines are that bird colonies should not be overflown below 2000 ft (∼610 m) above ground level and landings within 1/2 nautical mile (∼930 m) of bird colonies should be avoided wherever possible. These guidelines are less stringent and less specific than those that were recommended by the SCAR specialist group on birds, and represent a compromise to accommodate operational needs. While the adoption of clear and consistent guidelines for the operation of aircraft in Antarctica is welcome in that this provides practical advice that is likely to reduce incidences of close aircraft/bird encounters, there remains insufficient knowledge of the interactions between aircraft and birds in Antarctica, and the consequent impacts on individual birds and on bird populations. It is important, therefore, that the guidelines adopted are considered interim, and should be kept under scrutiny with revisions made as new and improved research results appear.     

The abundance of heterocystous blue-green algae in rice soils and inocula used for application in rice fields

Summary Algal populations were quantified (as colony-forming units [CFU] per square centimetre) in 102 samples of rice soils from the Philippines, India, Malaysia and Portugal, and in 22 samples of soil-based inocula from four countries. Heterocystous blue-green algae (BGA) were present in all samples. Nostoc was the dominant genus in most samples, followed by Anabaena and Calothrix. In soils, heterocystous BGA occurred at densities ranging from 1.0 × 10² to 8.0 × 10⁶ CFU/cm² (median 6.4 × 10⁴) and comprised, on average, 9% of the total CFU of algae. Their abundance was positively correlated with the pH and the available P content of the soils. In soil-based inocula, heterocystous BGA occurred at densities ranging from 4.6 × 10⁴ to 2.8 × 10⁷ CFU/g dw (dry weight), comprising only a moderate fraction (average 13%) of the total algae. In most soils, the density of indigenous N₂-fixing BGA was usually higher than that attained by applying recommended rates of soil-based inoculum. Whereas research on the practical utilization of BGA has been mostly directed towards inoculation with foreign strains, our results suggest that attention should also be given to agricultural practices that enhance the growth of indigenous strains already adapted to local environmental conditions.Maître de Recherches ORSTOM (France), Visiting Scientist at IRRI