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深海环境通常具有高盐,高压,高/低温,无光照等特点,使得海洋微生物存在一套独特的生理代谢机制和分子细胞结构,然而迄今绝大部分深海微生物不能在实验室条件下被分离培养,深海微生物资源开发遇到很大挑战。本研究通过不依赖培养的方法研究海洋微生物的基因资源,构建了南海深海沉积物fosmid宏基因组文库,共获得约39 600个克隆,插入片段范围在24~45 kb之间,平均插入片段大小为33 kb,克隆片段的总库容达到1 320 Mb。通过功能筛选获得3个具有淀粉酶活性的克隆子,选取其中最适温度较低的amy7作为进一步研究对象。构建amy7插入片段的重组质粒文库,获得一个同样有淀粉酶活性的克隆子amy7-6。经测序,克隆子amy7-6含有3 291 bp插入片段,序列比对分析后发现其中一个大小为1 920 bp的ORF,其编码的蛋白质序列(AmyS)与各种来源的糖苷酶有着较高的相似性。 相似文献
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采用化学裂解和酶解相结合的方法，进行了深海沉积物中微量DNA的提取，并采用DNA吸附树脂进行纯化。结果表明，该方法能有效地去除沉积物中的腐殖酸等抑制剂，每克湿重沉积物样品可得到DNA约16μg，回收率可达95％，所得到的DNA分子片段均在23kb左右，纯化后的DNA可直接应用于各种分子生物学操作。利用细菌16SrDNA通用引物对所提取的深海沉积物DNA进行了PCR—RRJ及系统发育分析，各主要细菌类群均能检出，证实该方法可以应用于深海等极端环境中微生物的多样性调查、系统发育分析以及特殊功能基因的筛选，同时还可应用于环境样品中生物量的半定量估计。 相似文献
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提取西太平洋"暖池"区深海沉积物样品总DNA,利用异化型亚硫酸盐还原酶(DSR)和甲基辅酶M还原酶(MCR)基因的特异引物,采用PCR-RFLP方法对沉积物样品中这两类功能基因多样性进行研究.该沉积物中的dsrAB基因分别来源于δ-变形菌中的6个属,其中最多的是脱硫弧菌属和脱硫杆状菌属;mcrA基因均来源于产甲烷古菌,其中主要是甲烷微菌.这些基因在系统发育树上都处于相对独立的分支.此外,还有较多的dsrAB,mcrA基因来源于未知的新属或新种.这些结果表明该海区沉积物中由微生物参与的硫和甲烷循环比较活跃,而且其中可能存在多种新的代谢途径. 相似文献
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北极深海沉积物中细菌和古菌群落结构研究 总被引：3，自引：0，他引：3
在北极深海沉积物生态系统中,微生物的群落结构由有机质输入、能量的可用性及其他环境因素决定.然而,全球气候变暖及其导致的冰盖提前融化正在影响微生物的多样性.为描述北极深海沉积物中的微生物群落结构及其与环境因素的相关性,我们利用罗氏454对北极深海沉积物样品的16S rDNA扩增子进行了测序,对细菌和古菌群落的丰富度、成分、结构及其系统发育分类地位进行了描述.硫还原和化能有机营养类是细菌群落中的主要类群;而古细菌群落主要是由微生物的关系最为密切的氨氧化奇古菌门(96.66%)和产甲烷古生菌界(3.21%).这项研究描述了北极极点附近深海沉积物(> 3500米)中的微生物多样性,将为以后研究类似环境中微生物代谢过程和途径等功能分析奠定基础. 相似文献
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《Deep Sea Research Part I: Oceanographic Research Papers》1999,46(6):1077-1094
In order to study temporal variations of the genetic material in the continental shelf and deep-sea sediments of the extremely oligotrophic Cretan Sea, samples were collected on seasonal basis from August 1994 to September 1995, with a multiple corer, at seven stations (from 40 to 1540 m depth). Surface sediments (0–1 cm) were sub-sampled and analyzed for nucleic acid content (DNA, RNA) and bacterial density. DNA concentrations in the sediments were high (on annual average, 25.0 μg g-1) and declined with increasing water depth, ranging from 3.5 to 55.2 μg g-1. DNA concentrations displayed wide temporal changes also at bathyal depths confirming the recent view of the large variability of the deep-sea environments. Also RNA concentrations decreased with increasing water depth (range: 0.4–29.9 μg g-1). The ratio of RNA to DNA did not show a clear spatial pattern but was characterized by significant changes between sampling periods. DNA concentrations were significantly correlated with protein and phytopigment concentrations in the sediment, indicating a possible relationship with the inputs of primary organic matter from the photic layer. Bacterial densities were generally high (range: 0.9–4.6×108 cells g-1) compared to other deep-sea environments and decreased with increasing water depth. Estimates of the bacterial contribution to the sedimentary genetic material indicated that bacterial-DNA accounted, on annual average, for a small fraction of the total DNA pool (4.3%) but that bacterial-RNA represented a significant fraction of the total sedimentary RNA (26%). Bacterial contribution to nucleic acids increased, even though irregularly, with increasing depth. In deep-sea sediments, changes in RNA concentrations appear to be largely dependent upon bacterial dynamics. Estimates of the overall living contribution to the DNA pools (i.e. microbial plus meiofaunal DNA) indicated that the large majority (about 90%) of the DNA in continental and deep-sea sediments of the eastern Mediterranean was detrital. The non-living DNA pools reach extremely high concentrations up to 0.41 g DNA m-2 cm-1. Thus, especially in deep benthic habitats, characterized by low inputs of labile organic compounds, detrital DNA could represent a suitable and high quality food source or a significant reservoir of nucleic acid precursors for benthic metabolism. 相似文献
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环境样品的低生物量是微生物宏基因组学研究面临的首要挑战,通过基因组扩增技术来满足高通量测序对DNA样品量的需求是最常用的解决策略。MALBAC(Multiple Annealing and Looping Based Amplification Cycles)基因组扩增试剂盒最初为扩增和研究哺乳动物的单细胞基因组而研发。本文中,我们通过人工构建的微生物群落来检测该试剂盒在微生物宏基因组扩增方面的效率和应用可行性。结果表明,每个标准反应中,10 pg的DNA模板量足以满足MALBAC试剂盒对样品扩增的需要。每个标准反应DNA模板用量为10和100 pg时,所扩增DNA样品的基因组覆盖度与原始未扩增样品表现出高度的一致性,证明MALBAC试剂盒扩增效果的高度稳定性和一致性。常用的GenomePlex全基因组扩增试剂盒使我们可以在每个标准反应DNA模板量为100 pg的条件下扩增获得足够的DNA样品,但是结果表明该参照试剂盒无法有效的实现对群落中低丰度细菌菌株基因组的线性扩增。对于MALBAC试剂盒和参照试剂盒而言,在扩增高GC含量的微生物物种基因组DNA方面效率低下。我们的实验结果表明MALBAC试剂盒在高效扩增环境样品宏基因组DNA方面的可行性,但对该试剂盒在扩增环境样品中高GC含量微生物物种方面的适用性存在疑虑。 相似文献
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AbstractDue to the low shear strength of deep-sea sediment, crawled mineral collector is easily slips in the deep-sea mining process, therefore, high-traction bionic grouser is needed to be studied to improve the working efficiency. Based on the rate-dependent characteristics of deep-sea sediment, a rate-dependent extended Drucker-Prager material constitutive model is used to define the deep-sea sediment. The Arbitrary Lagrange-Euler finite element (LEA-FE) was used to simulate the cutting process of different bionic grousers at different speeds. By comparing the simulation of different grousers, it was found that the maximum traction of grouser is related to the grouser parameters (distance L from the top to the curvature change point and curvature radius R). By analyzing the traction characteristics of different bionic grousers, the binary quadratic regression equation between maximum traction and bionic grouser parameters was established and the best bionic grouser parameters were obtained by the optimization algorithm. Based on the rate-dependent properties of deep-sea sediment, the traction characteristics at different speeds were analyzed and the relationship between maximum traction and speeds was established, the best bionic grouser walking speed was obtained, which can provide the theoretical basis for the crawled mineral collector. 相似文献
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S. Vanucci A. Dell'Anno A. Pusceddu M. Fabiano R. S. Lampitt R. Danovaro 《Progress in Oceanography》2001,50(1-4)
Downward fluxes of microbial assemblages associated with sinking particles sampled in sediment traps deployed at nominal depths of 1000 m (trap A), 3000 m (trap B) and 4700 m (trap C) were measured between October 1995 and August 1998 on the Porcupine Abyssal Plain (PAP, NE Atlantic). The goal of the study was to provide detailed information on the microbial contributions to the particulate organic carbon and DNA fluxes. Bacterial fluxes associated with settling particles in the PAP area were generally low and significantly lower than bacterial fluxes reported from the same area during 1989–90. Marked seasonal pulses in the microbial assemblages were observed in all years that were associated with particle flux maxima in April–June. No significant differences were found in microbial fluxes between 1000 and 4700 m depth, but both the bacterial biomass flux and the frequency of dividing bacteria increased with depth, suggesting that organic matter turnover and conversion into bacterial biomass increased in the deeper traps. The structure of microbial assemblages displayed clear changes with increasing depth; the ratios of bacteria to both flagellates and cyanobacteria increased up to 4-fold between 1000 and 4700 m, showing a marked increase in bacterial dominance in the deeper layers of the water column. A parallel increase of the bacterial contribution to particulate organic carbon (POC) and DNA fluxes was observed. Total microbial contribution to the POC flux in the PAP area was about 2%, whereas the contribution of cyanobacteria was negligible. Fluxes of microbial assemblages were significantly correlated with DNA fluxes and on average the bacteria accounted for 5% of DNA fluxes. Data reported here confirm that the “rain” of particulate bacterial DNA may represent an important source of nucleotides for deep-sea bacteria, but also suggests that a much larger pool of detrital DNA is potentially available to deep-sea micro-organisms. 相似文献
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Ｄｅｔｅｃｔｉｏｎ ｏｆ Ｈｅｐａｔｉｔｉｓ Ａ ｖｉｒｕｓ ｉｎ ｓｈｅｌｌｆｉｓｈ ｂｙ ｒｖｅｒｓｅ ｔｒａｎｓｃｒｉｐｔｉｏｎ ＰＣＲTXＤｅｔｅｃｔｉｏｎｏｆＨｅｐａｔｉｔｉｓＡｖｉｒｕｓｉｎｓｈｅｌｌｆｉｓｈｂｙｒｅｖｅｒｓｅｔｒａｎｓｃｒ... 相似文献
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Namirimu Teddy Park Mi-Jeong Kim Yun Jae Lim Dhongil Lee Jung-Hyun Shin Ayoung Kim Dongsung Kwon Kae Kyoung 《Ocean Science Journal》2023,58(2):1-15
Since the discovery of hydrothermal vents in the late 1970s, deep-sea hydrothermal vent fields have attracted great attention as biological hotspots. However, compared with other ocean ridges, the structure and function of microbial communities inhabiting vent fields in the Central Indian ridge (CIR) remain understudied. Here, we provide for the first time 16S rRNA gene-based comparative metagenomic analysis of the sediment-associated microbial communities from three newly discovered vent fields in the CIR. Sediment samples collected in the Invent B, Invent E and Onnuri vent fields varied in geochemical properties, elemental concentrations and associated microbial communities. Proteobacteria (Gammaproteobacteria) was the dominant phylum in Invent B and Onnuri vent fields. In contrast, Invent E mainly consisted of Chloroflexi and Euryarchaeota. Predicted functional profiling revealed that the microbial communities in the three vents are dominated by chemoheterotrophic functions. In addition, microbial communities capable of respiration of sulfur compounds, nitrification, nitrite oxidation, methylotrophy, and methanotropy were found to be the main chemolithoautotrophs. Compared to other vent fields, Invent E showed a predominance of archaeal methanogens suggesting it exhibits slightly different geochemistry. Multivariate analysis indicated that the biogeochemical and trace metal differences are reflected in the sediment microbial compositions of the three vent fields. This study expands our current understanding of the microbial community structure and potential ecological functions of the newly discovered hydrothermal vent fields in the CIR.
相似文献14.
WEI Manman ZHANG Rubing WANG Yuguang JI Houguo ZHENG Ji CHEN Xinhu ZHOU Hongbo 《海洋学报(英文版)》2013,32(2):42-51
The aim of this study is to investigate microbial structures and diversities in five active hydrothermal fields’ sediments along the Eastern Lau Spreading Centre (ELSC) in the Lau Basin (southwest Pacific). Microbial communities were surveyed by denatured gradient gel electrophoresis (DGGE) and clone library analysis of 16S rRNA genes. The differences in microbial community structures among sediment samples from the five deep-sea hydrothermal sites were revealed by DGGE profiles. Cluster analysis of DGGE profiles separated the five hydrothermal samples into two groups. Four different 16S rRNA gene clone libraries, representing two selected hydrothermal samples (19-4TVG8 and 19-4TVG11), were constructed. Twenty-three and 32 phylotypes were identified from 166 and 160 bacterial clones respectively, including Proteobacteria, Bacteroidetes, Firmicutes, Nitrospirae and Planctomycetes. The phylum Proteobacteria is dominant in both bacterial libraries with a predominance of Gamma-Proteobacteria. A total of 31 and 25 phylotypes were obtained from 160 and 130 archaeal clones respectively, including Miscellaneous Crenarchaeotic Group, Marine Group Ⅰ and Ⅲ, Marine Benthic Group E, Terrestrial Hot Spring Crenarchaeota and Deep-sea Hydrothermal Vent Euryarchaeota. These results show a variety of clones related to those involved in sulfur cycling, suggesting that the cycling and utilization of sulfur compounds may extensively occur in the Lau Basin deep-sea hydrothermal ecosystem. 相似文献
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富钴结壳是一种重要的海底矿产资源.富钴结壳是生物地球化学和微生物学的研究热点,但目前我们对富钴结壳区的微生物群落结构仍缺乏认识.本研究针对中太平洋海山富钴结壳区沉积物,采用非培养方法获得微生物基因组信息并分析其生态学功能.采用PCR的方法筛选得到78个带有16S rRNA基因的克隆.变形杆菌门和奇古菌门MGI类群分别为古菌和细菌的主要类群.测序并分析了9个带有16S rRNA 基因的fosmid克隆.这些克隆中含有较多的代谢相关和重金属抗性相关基因,表明多样的代谢和抗逆途径在微生物适应富钴结壳环境中发挥了重要作用.代谢途径分析表明,氮循环是富钴结壳区一种重要的生物地球化学过程.此外,发现多个水平基因转移事件,这些基因大部分转移自细菌,少量转移自古菌或植物.比较基因组分析表明,属于奇古菌门MGI类群的克隆W4-93包含一段共线性的基因簇.以上结果说明水平基因转移和基因组分歧进化在微生物适应深海环境过程中发挥了重要作用. 相似文献
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以萘为唯一碳源和能源从南，中绳海槽深海沉积物中分离得到一株能降解萘的海洋嗜低温细菌Nah-1，测定了该茵的最适生长条件及生长曲线。16S rRNA基因（16SrDNA）序列同源性分析表明该菌属于解环菌属（Cyczocznsticus）。PCR扩增萘降解基因得到目标片段，比对结果表明，相似度最高的基因phnAl来自Cycloclasticus sp．A5，为99％，该基因编码的蛋白是萘双加氧酶大亚基。 相似文献
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DNA的简便高效制备方法是研究基因结构、功能及开展其它各项研究的基础。首次报道了针对节旋藻的结构和组成特性而建立的制备节旋藻高分子量DNA的两种方法。其中第一个方法是常规方法，可大量提取纯度较高、分子量达50kb的节旋藻DNA，可用于构建节旋藻质粒库、Southern杂交和PCR操作等；第2种方法是用脉冲电场凝胶电泳分离DNA片段，制备的DNA片段达数百kb，可用于构建节旋藻噬菌体库、粘粒库、BAC库等，从而为构建节旋藻物理图谱，定位克隆基因奠定基础。 相似文献
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东太平洋深海沉积物中DNA的提取及细菌多样性初步分析 总被引：1，自引：0，他引：1
以东太平洋海隆附近深海柱状沉积物为材料，通过化学裂解和酶消化相结合的方法提取了沉积物微生物的总基因组DNA，并进行了纯化。结果表明所得到的DNA分子片段大小在21kb左右，纯化后的DNA可直接用于PCR等分子生物学操作。细菌16SrDNAV3可变区的PCR—DGGE图谱展示出15条以上条带，表明深海沉积物中细菌多样性较高，群落结构比较复杂。对其中9条主要条带进行回收、测序和系统发育分析，结果表明所获得的序列分属放线菌门（Actinobacteria），绿弯菌门（Chloroflexi），γ-变形细菌亚门（Gamma—proteobacteria），α-变形细菌亚门（Alpha—proteobacteria）和嗜酸菌门（Acidobacteria）5个大类群。 相似文献
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海底底质的物理力学参数不同于陆地土壤,其极低的抗剪强度和承压强度对深海采矿车的行走性能提出高要求。分析基于车辆地面力学理论,开展了底质土力学特性试验研究,建立了深海底质力学模型。根据深海重载作业采矿车样机结构参数,在大型动力学仿真软件Recurdyn中建立了仿真模型。通过直线行走多体动力学仿真与直线行走海试试验的对比,验证了仿真模型的准确性。在此基础上开展了采矿车样机在深海软底质上的多种行走工况动力学仿真,分析与评价其行走性能。结果表明,采矿车样机可以顺利完成转弯、爬坡、越障等基本功能。该研究成果可为深海采矿车海底行走性能评估提供理论参考,为深海采矿车和软底质的相互作用力学研究提供借鉴。 相似文献
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Jenny Brunnegrd Sibylle Grandel Henrik Sthl Anders Tengberg Per O.J. Hall 《Progress in Oceanography》2004,63(4):159-181
Rates of transformation, recycling and burial of nitrogen and their temporal and spatial variability were investigated in deep-sea sediments of the Porcupine Abyssal Plain (PAP), NE Atlantic during eight cruises from 1996 to 2000. Benthic fluxes of ammonium (NH4) and nitrate (NO3) were measured in situ using a benthic lander. Fluxes of dissolved organic nitrogen (DON) and denitrification rates were calculated from pore water profiles of DON and NO3, respectively. Burial of nitrogen was calculated from down core profiles of nitrogen in the solid phase together with 14C-based sediment accumulation rates and dry bulk density. Average NH4 and NO3-effluxes were 7.4 ± 19 μmol m−2 d−1 (n = 7) and 52 ± 30 μmol m−2 d−1 (n = 14), respectively, during the period 1996–2000. During the same period, the DON-flux was 11 ± 5.6 μmol m−2 d−1 (n = 5) and the denitrification rate was 5.1 ± 3.0 μmol m−2 d−1 (n = 22). Temporal and spatial variations were only found in the benthic NO3 fluxes. The average burial rate was 4.6 ± 0.9 μmol m−2 d−1. On average over the sampling period, the recycling efficiency of the PON input to the sediment was 94% and the burial efficiency hence 6%. The DON flux constituted 14% of the nitrogen recycled, and it was of similar magnitude as the sum of burial and denitrification. By assuming the PAP is representative of all deep-sea areas, rates of denitrification, burial and DON efflux were extrapolated to the total area of the deep-sea floor (>2000 m) and integrated values of denitrification and burial of 8 ± 5 and 7 ± 1 Tg N year−1, respectively, were obtained. This value of total deep-sea sediment denitrification corresponds to 3–12% of the global ocean benthic denitrification. Burial in deep-sea sediments makes up at least 25% of the global ocean nitrogen burial. The integrated DON flux from the deep-sea floor is comparable in magnitude to a reported global riverine input of DON suggesting that deep-sea sediments constitute an important source of DON to the world ocean. 相似文献