分子生态学方法对油藏细菌群落结构分析的比较

目前对油藏细菌群落结构分析方法繁多,为得到准确全面的群落分析结果,必须选取合适的方法。群落分析方法的优劣进行比较分析,选取从新疆陆梁油田注水井筛出的11株单菌,测定其16srDNA序列,制成包含7个菌属、9个菌种的混合菌液。应用基于PCR扩增的三种分子生态技术DGGE、T-RFLP、建立16SrDNA文库比较和分析了混合菌液细菌多样性。使用PCR-DGGE方法时发现,DGGE方法可灵敏地检测到序列1 bp的变化,能将不同菌株分开,但该方法经过切胶测序后的的目标序列较短,信息量不大且有时有一定误差,较适合用于定性对比;而用于菌群结构的分析时应结合其他方法。T-RFLP法可区分大部分菌属,但数据库不够完整,不能确定细菌群落组成;因此也不适合单独用在细菌群落检测中,可用作多个样品种群多样性的对比或结合其他方法对样品进行系统透彻解析。建立16srDNA克隆文库法成功鉴定到7个菌属8个菌种16SrDNA序列,更适用于油藏细菌群落结构的分析。     

Chloroplast DNA sequence from a miocene Magnolia species

DNA has been successfully extracted from several samples of preserved tissue, the oldest so far reported originating from a 13,000-year-old ground sloth. Both severe damage to the preserved DNA, primarily due to oxidation of the pyrimidines, has prevented the acquisition of sequence data from ancient samples except in a few cases. We report here the extraction of DNA from fossil leaf samples from the Miocene Clarkia deposit (17-20 Myr old), the amplification of an 820-base pair (bp) DNA fragment from the chloroplast gene rbcL from a fossil of the genus Magnolia, and its subsequent sequencing. The sequence was verified by comparison with published and unpublished rbcL sequences. These results extend our ability to analyse ancient DNA and may open new avenues into problems in palaeobotany, biogeography, and in the calibration of mutation rates.     

Isolation of a 250 million-year-old halotolerant bacterium from a primary salt crystal

Bacteria have been found associated with a variety of ancient samples, however few studies are generally accepted due to questions about sample quality and contamination. When Cano and Borucki isolated a strain of Bacillus sphaericus from an extinct bee trapped in 25-30 million-year-old amber, careful sample selection and stringent sterilization techniques were the keys to acceptance. Here we report the isolation and growth of a previously unrecognized spore-forming bacterium (Bacillus species, designated 2-9-3) from a brine inclusion within a 250 million-year-old salt crystal from the Permian Salado Formation. Complete gene sequences of the 16S ribosomal DNA show that the organism is part of the lineage of Bacillus marismortui and Virgibacillus pantothenticus. Delicate crystal structures and sedimentary features indicate the salt has not recrystallized since formation. Samples were rejected if brine inclusions showed physical signs of possible contamination. Surfaces of salt crystal samples were sterilized with strong alkali and acid before extracting brines from inclusions. Sterilization procedures reduce the probability of contamination to less than 1 in 10(9).     

东北梅花鹿肠道纤维素降解微生物的分离筛选

食草动物依赖胃肠道微生物分解利用天然木质纤维素物质。为获得高效纤维素降解菌，本文将东北梅花鹿粪便样品，经过研磨，重悬于羧甲基纤维素钠（CMC-Na）作为唯一碳源的富集培养基中，进行富集培养和筛选，用革兰氏碘液染色显示纤维素水解圈，通过纤维素水解圈直径（D）和菌落直径（d）比值，初步判断纤维素降解菌降解纤维素能力．筛选出106株纤维素降解微生物，包括72株细菌、22株真菌和12株放线菌．细菌16S rDNA和真菌ITS序列测序结果显示，这些纤维素降解菌主要集中分布于变形菌门（Proteobacteria）、厚壁菌门（Firmicutes）、放线菌门（Actinobacteria）和子囊菌门（Ascomycota）．本文提供了一种快速有效的反刍动物肠道纤维素降解微生物的筛选方法，为东北梅花鹿肠道微生物高通量测序结果提供实验数据支持．      

Constraint on the late Mesozoic regional angular unconformity in West Liaoning--North Hebei by LA-ICP-MS dating

The angular unconformity between the Tuchengzi Formation and the Zhangjiakou Formation in the Chengde area is commonly regarded as the direct evidence of boundary between Jurassic and Cretaceous systems, and as the evidence of tectonic transformation interfaces in Mesozoic Era in the West Liaoning-North Hebei. Moreover, the lower boundary age of the Zhangjiakou Formation in the Chengde was to be thought about 135 Myr in the past. Based on the detailed study and systematic sampling in Yixian-Beipiao, Lingy- uan, Chengde and Luanping areas in the field, the authors obtained the following isotopic ages by means of LA-ICP-MS dating： In the Lingyuan area, the Zhangjiakou Formation and the Yixian Formation are in angular unconformable contact; the age range of the Zhangjiakou Formation volcanic rock is from 133 to 130 Myr （seven samples）, and that of the Yixian Formation volcanic rock is from 126 to 121 Myr （eleven samples）. In the Luotuoshan of Chengde, the Tuchengzi Formation and the Zhangjiakou Formation are in angu- lar unconformable contact; the age at the lower Zhangjiakou Formation is about 124 Myr （one sample）, which reflects that the age range of the lower Zhangjiakou Formation is from 129 to 124 Myr combined with the previous study. In the Luanping basin, there is the larg- est sediment gap in upper Mesozoic, whose lower boundary age is confined in 131 to 130 Myr. In the Yixian-Beipiao area, the second Member of the Yixian Formation, making up more than 3/4 thickness of the Yixian Formation, was formed in 126 to 119 Myr, and the age of the Yixian Formation at the bottom is about 132 Myr. Moreover, the upper boundary age of the Tuchengzi Formation is about 136 Myr in the West Liaoning-North Hebei region. Based on the above isotopic data combined with the geological characteristics in the field in the West Liaoning-North Hebei region, it reflects that there was a widespread tectonic event in the period of 136 to 126 Myr （mainly in 130-126 Myr） in the West Liaoning-North Hebei, and it formed th     

不同保存方式下金龟甲虫DNA提取方法及28S rDNA序列分析

通过电泳检测和PCR扩增28SrDNA核基因序列的结果,比较了无水乙醇、75%乙醇、针插干制3种金龟甲虫标本的保存方式和SDS-蛋白酶K法、CTAB法、饱和NaCl法3种提取DNA方法.结果显示,SDS-蛋白酶K法、CTAB法提取3种保存方式的标本时,可成功提取DNA并有效扩增目的基因;饱和NaCl法仅能提取无水乙醇保存标本的DNA并扩增目的基因,而75%乙醇保存标本和干制标本的DNA提取和基因扩增均不理想.     

LA-ICP-MS dating of Zhangjiakou Formation volcanic rocks in the Zhangjiakou region and its geological significance

The geological cross section of the Zhangjiakou Formation is originally established in the Zhangjiakou region. From the dating of four samples from typical cross sections of the Zhangjiakou Formation in the Zhangjiakou region, the age range is found to be mainly from 143.0 ± 3.7 Myr to 136.1 ± 1.4 Myr, which almost equals the sum of the age range of the Zhangjiakou Formation （136-135 Myr） and that of the Tuchengzi Formation （143-136 Myr） in the Luanping region, and the Zhangjiakou and Tuchengzi Formations in Zhang- jiakou-Luanping of North Hebei are found to be conformable. The evidence above indicates that the Zhangjiakou and Tuchengzi Formations, which are distributed in Zhangjiakou-Luanping, were formed under the same geological setting. Volcanic activity was earlier and longer （143-136 Myr） in the western region （Zhangjiakou） than that in the eastern region （Luanping）. The ＂Zhangjiakou Formation＂ in Chengde and Pingquan formed in the period in which the Yixian Formation in West Liaoning formed, so should be renamed the Yixian Formation.     

Diversity of 16S rDNA and environmental factor influencing microorganisms in Malan ice core

The research on extrempholic microorganisms in glacial low-temperature environment receives more attention than ever before. Due to the successive chronological records in ice core, it is important to initiate microbiological studies on ice core samples. 23 samples from one ice core,drilled from central Qinghai-Tibetan Plateau, were analyzed.The number of total microorganisms and culturable microorganisms in different layers showed that it related with the content of dust in ice. It is suggested that the distribution of microorganisms in ice depends on the transportation of materials during glacier development. The bacteria diversity in Malan Glacier was analyzed by 16S rDNA sequencing methods, which showed that many sequences were similar to known psychrophilic bacteria.     

Constraints on the late Mesozoic regional angular unconformity in West Liaoning-North Hebei by LA-ICP-MS dating

The angular unconformity between the Tuchengzi Formation and the Zhangjiakou Formation in the Chengde area is commonly regarded as the direct evidence of boundary between Jurassic and Cretaceous systems, and as the evidence of tectonic transformation interfaces in Mesozoic Era in the West Liaoning-North Hebei. Moreover, the lower boundary age of the Zhangjiakou Formation in the Chengde was to be thought about 135 Myr in the past. Based on the detailed study and systematic sampling in Yixian-Beipiao, Lingyuan, Chengde and Luanping areas in the field, the authors obtained the following isotopic ages by means of LA-ICP-MS dating: In the Lingyuan area, the Zhangjiakou Formation and the Yixian Formation are in angular unconformable contact; the age range of the Zhangjiakou Formation volcanic rock is from 133 to 130 Myr (seven samples), and that of the Yixian Formation volcanic rock is from 126 to 121 Myr (eleven samples). In the Luotuoshan of Chengde, the Tuchengzi Formation and the Zhangjiakou Formation are in angular unconformable contact; the age at the lower Zhangiiakou Formation is about 124 Myr (one sample), which reflects that the age range of the lower Zhangjiakou Formation is from 129 to 124 Myr combined with the previous study. In the Luanping basin, there is the largest sediment gap in upper Mesozoic, whose lower boundary age is confined in 131 to 130 Myr. In the Yixian-Beipiao area, the second Member of the Yixian Formation, making up more than 3/4 thickness of the Yixian Formation, was formed in 126 to 119 Myr, and the age of the Yixian Formation at the bottom is about 132 Myr. Moreover, the upper boundary age of the Tuchengzi Formation is about 136 Myr in the West Liaoning-North Hebei region. Based on the above isotopic data combined with the geological characteristics in the field in the West Liaoning-North Hebei region, it reflects that there was a widespread tectonic event in the period of 136 to 126 Myr (mainly in 130-126 Myr) in the West Liaoning-North Hebei, and it formed the late Mesozoic regional angular unconformity. Moreover, the lower boundary ages of the late Mesozoic regional angular unconformity in different areas are different, and the tectonics and the creature assemblage are different in the strata at the top and bottom of the regional angular unconformity. It implies that the late Mesozoic regional angular unconformity is a regional interface of tectonic transformation in Mesozoic Era in the West Liaoning-North Hebei.     

四川盆地海相三叠纪硫同位素组成与地质成因分析

文章通过对四川盆地海相三叠纪蒸发岩硫同位素δ34S组成特征的对比分析,针对该区域三叠系δ34S具有异常高值并且呈现随时间由早到晚逐渐轻化的特点,结合其沉积地质背景、物化条件,特别是对复杂条件下同位素分馏、地质地球化学特征的研究,对硬石膏的δ34S异常高值和随蒸发而逐渐贫化的地质成因进行了探讨,旨在为重建古环境及地层的划分和对比提供重要依据。研究表明,硬石膏中δ34S异常高值的产生主要是处在封闭环境、硫酸盐细菌还原时的同位素分馏作用的结果。     

Diversity of 16S rDNA and environmental factor influncing microorganisms in Malan ice core

The research on extrempholic microorganisms in glacial low-temperature environment receives more attention than ever before. Due to the successive chronological records in ice core, it is important to initiate microbiological studies on ice core samples. 23 samples from one ice core, drilled from central Qinghai-Tibetan Plateau, were analyzed. The number of total microorganisms and culturable microorganisms in different layers showed that it related with the content of dust in ice. It is suggested that the distribution of microorganisms in ice depends on the transportation of materials during glacier development. The bacteria diversity in Malan Glacier was analyzed by 16S rDNA sequencing methods, which showed that many sequences were similar to known psychrophilic bacteria.     

一株具抗肿瘤活性的北极细菌的筛选及分子鉴定

在中国首次北极科学考察期间，从北极海域采集了水体，沉积物等样品，从中分离得到一批嗜冷细菌，采用MTT法对这些北极海洋细菌进行了细胞毒活性物质的筛选，得到一株具有细胞毒活性的菌株。采用16SrDNA序列分析及系统发育分析方法对该株菌株进行了分子鉴定。     

寒武系盐盆地的分布特征及其对中国成盐找钾的意义

 寒武系膏盐岩在世界范围内广泛分布,以亚洲寒武系分布最为广泛,含盐岩系主要分布在俄罗斯东西伯利亚、伊朗—印度—巴基斯坦—阿曼和中国.东西伯利亚、伊朗—印度—巴基斯坦均有钾盐发现.构造基底性质对钾盐盆地的形成具有关键性控制作用：稳定构造区在准稳定区有利聚钾;准稳定区在局部稳定构造区利于聚盐成钾,如形成于早寒武世的涅帕钾盐矿床——世界上大型钾盐矿之一,即位于前寒武纪地层为基底的古拗陷区.中国主要的古代盐盆地多发育于以前寒武纪为基底的准克拉通(陆块)中.近年来,随着油气勘查和盐业开发工作,中国上扬子陆块、华北陆块和塔里木陆块相继发现了4个大型的膏盐岩盆地.上扬子陆块与塔里木陆块发现单层厚度较大的岩盐沉积及较好的成钾水化学显示,为在中国寒武系蒸发岩盆地开展成盐找钾工作提供了重要依据.     

新疆达坂城盐湖中度嗜盐菌的16SrDNA序列研究

中度嗜盐菌作为一类微生物资源,已经在很多方面应用。从新疆达坂城盐湖样品中分离得到17株中度嗜盐菌。其中11株为革兰氏阳性,6株为革兰氏阴性,并完成表型和16SrDNA序列的测定．其表塑特征和16SrDNA序列分析结果表明这些菌分别属于Halomonas、Bacillus、Salinicoccus、Halobacillus、Marinococcus、Thalassobacillus、Nesterenkonia属,其中大部分属于Halomonas属。     

一种河口沉积物总DNA的高效提取方法

研究主要目的是从河口沉积物中提取高质量DNA,为后期应用分子生物学技术研究河口沉积物微生物功能基因及种群分析等奠定基础。用CTAB-SDS-冻融法,样品预处理-CTAB-SDS-冻融法,样品预处理-CTABSDS-冻融-DNA重沉淀法和土壤总DNA提取试剂盒法提取3种河口沉积物总DNA,通过DNA琼脂糖凝胶电泳、DNA纯度和浓度及16S rDNA的PCR扩增分别对这4种方法进行评价。结果显示,样品预处理-CTABSDS-冻融法-DNA重沉淀所提取的DNA质量最高,可以用于后续的分子生物学研究。     

Change of bacterial community in the Malan Ice Core and its relation to climate and environment

In order to understand the relationship between the community structure of bacteria in ice core and the past climate and environment, we initiated the study on the microorganisms in the three selected ice samples from the Malan ice core drilled from the Tibetan Plateau. The 16S ribosomal DNA (rDNA) molecules were directly amplified from the melt water samples, and three 16S rDNA clone libraries were established. Among 94 positive clones, eleven clones with unique restriction pattern were used for partial sequence and compared with eight reported sequences from the same ice core. The phylotypes were divided into 5 groups: alpha, beta, gamma proteobacteria; CFB, and other eubacteria group. Among them, there were many “typical Malan glacial bacteria“ pertaining to psychrophilies and new bacteria found in the ice core. At a longer time scale, the concentration distribution of “typical Malan glacial bacteria“ with depth showed negative correlation with temperature variations and was coincident with dirty layer. It implied the influence of temperature on the microbial record through impact on the concentrations of the “typical Malan glacial bacteria“. In addition, the nutrition contained in ice was another important factor controlling the distribution of microbial population in ice core section. Moreover, the result displayed an apparent layer distribution of bacterial community in the ice core section, which reflected the microbial response to the past climatic and environmental conditions at the time of deposition.     

分子生物学技术在微生物多样性研究中的应用

用传统的方法很难鉴别微生物的差异,分子生物学技术就很好地解决了该问题,尤其适合于常规方法失效的情况下.该技术的主要程序是:先从微生物中直接提取DNA,对DNA进行PCR(PolymericChain Reaction)或RAPD(Random Amplification of Polymorphic DNA)扩增,然后进行DGGE(DenaturingGradient Get Electrophoresis)电泳,或RFLP(Restriction Fragment Lergth Polymorphism),ELISA(Enzyme-Linked Immunosorbent Assav)分析等,之后进行测序,测序结果与网上基因库进行对比,来确定样品中微生物的种类.此方法是直接从样品中提取总DNA,中间不会造成菌株的富集或衰减,具有很高的准确性.此方法对微生物多样性的深入研究很有效.     

新疆泥火山群土壤细菌群落的DGGE分析

采用变性梯度凝胶电泳(DGGE)指纹图谱技术,对新疆乌苏泥火山区土壤细菌群落的16S rDNA V3区进行电泳分离,并用Quantity one软件对图谱进行聚类分析,对主要条带进行回收测序.DGGE图谱显示,各土样细菌的组成有显著差异.聚类分析表明,相同月份、不同深度土样细菌组成相近.特异条带的回收测序结果表明,假单胞菌属是泥火山土壤优势菌群.新疆泥火山区土壤细菌多态性明显,并容易受生态、气候等因素影响,但优势菌群受此影响较小.     

Atmospheric carbon dioxide concentrations over the past 60 million years

Knowledge of the evolution of atmospheric carbon dioxide concentrations throughout the Earth's history is important for a reconstruction of the links between climate and radiative forcing of the Earth's surface temperatures. Although atmospheric carbon dioxide concentrations in the early Cenozoic era (about 60 Myr ago) are widely believed to have been higher than at present, there is disagreement regarding the exact carbon dioxide levels, the timing of the decline and the mechanisms that are most important for the control of CO2 concentrations over geological timescales. Here we use the boron-isotope ratios of ancient planktonic foraminifer shells to estimate the pH of surface-layer sea water throughout the past 60 million years, which can be used to reconstruct atmospheric CO2 concentrations. We estimate CO2 concentrations of more than 2,000 p.p.m. for the late Palaeocene and earliest Eocene periods (from about 60 to 52 Myr ago), and find an erratic decline between 55 and 40 Myr ago that may have been caused by reduced CO2 outgassing from ocean ridges, volcanoes and metamorphic belts and increased carbon burial. Since the early Miocene (about 24 Myr ago), atmospheric CO2 concentrations appear to have remained below 500 p.p.m. and were more stable than before, although transient intervals of CO2 reduction may have occurred during periods of rapid cooling approximately 15 and 3 Myr ago.     

油页岩微生物总DNA的提取方法

为建立油页岩微生物总DNA提取方法,以两矿区的6个样品为材料,采用SDS高盐、SDS液氮研磨、SDS反复冻融、SDS异硫氰酸胍和试剂盒提取法分别对总DNA进行提取.结果表明,各方法提取效果差异很大,改进的SDS高盐提取法效果最好,各样品均得到约23kb较完整片段,且产率显著高于其他方法,达9144~38685ng·g-1干样;以未纯化的DNA为模板,对16SrDNA进行PCR-DGGE,得到了相应的扩增产物及DGGE指纹图谱,说明该法适合油页岩样品.SDS液氮研磨和SDS反复冻融提取法仅能得到部分样品总DNA,且产率和纯度较低,而SDS异硫氰酸胍和试剂盒提取法无DNA提出,不适合此类样品.