A bacterium that can grow by using arsenic instead of phosphorus

A bacterium that can grow by using arsenic instead of phosphorus

一株可能利用砷替代磷作為生長所需元素的細菌

ABSTRACT:

Life is mostly composed of the elements carbon, hydrogen, nitrogen, oxygen, sulfur and phosphorus. Although these six elements make up nucleic acids, proteins and lipids and thus the bulk of living matter, it is theoretically possible that some other elements in the periodic table could serve the same functions. Here we describe a bacterium, strain GFAJ-1 of the Halomonadaceae, isolated from Mono Lake, CA, which substitutes arsenic for phosphorus to sustain its growth. Our data show evidence for arsenate in macromolecules that normally contain phosphate, most notably nucleic acids and proteins. Exchange of one of the major bio-elements may have profound evolutionary and geochemical significance.

生命主要由碳，氫，氮，氧，硫和磷等元素所組成,然而這六種元素可組成核酸,蛋白質和脂質,因此這些大量的生命組成物質,理論上在元素週期表上可能會有其他元素有相同的功能.在此我們描述一株Halomonadaceae屬的細菌strain GFAJ-1，分離自加州的Mono湖.一種利用砷替代磷作為生長所需元素的細菌.研究數據顯示在正常含有磷酸鹽的大分子中含有砷酸鹽,尤其是核酸和蛋白質.替換主要的生物元素可能會深深影響演化和地球化學的重要性.

Biological dependence on the six major nutrient elements carbon, hydrogen, nitrogen, oxygen, sulfur, and phosphorus is complemented by a selected array of other elements, usually metal(loid)s present in trace quantities that serve critical cellular functions, such as enzyme cofactors(1). There are many cases of these trace elements substituting for one another. A few examples include the substitution of tungsten for molybdenum and cadmium for zinc in some enzyme families (2, 3) and copper for iron as an oxygen-carrier in some arthropods and mollusks(4). In these examples and others, the trace elements that interchange share chemical similarities that facilitate the swap. However, there are no prior reports of substitutions for any of the six major

elements essential for life. Here we present evidence that arsenic can substitute for phosphorus in the biomolecules of a naturally-occurring bacterium.

生物學證明6種主要營養元素碳，氫，氮，氧，硫和磷的補充是藉由從其他元素中經一系列選擇的結果，通常微量金屬的存在可提供一些重要的細胞功能.例如作為酵素輔因子.這些微量元素間彼此替換的例子很多,一些例子包含:在一些酵素家族中(2，3)用鎢代替鉬和用鎘代替鋅. 以及在一些節足動物和軟體動物中利用銅代替鐵做為氧氣輸送的角色. 在這些例子及其他例子中,微量元素的替換,均分了化學性質的相似性並促進了替換。然而先前的研究報告中,還沒有任何一篇有證實,在構成生命體的六種主要元素中有任何的可替代元素出現.在此研究中證實了於自然界存在的細菌生物分子中砷可以替代磷.

Arsenic (As) is a chemical analog of phosphorus (P), which lies directly below P on the periodic table. Arsenic possesses a similar atomic radius, as well as near identical electronegativity to P(5). The most common form of P in biology is phosphate (PO43-), which behaves similarly to arsenate (AsO43-) over the range of biologically relevant pH and redox gradients (6). The physico-chemical similarity between AsO43-and PO43-contributes to the biological toxicity of AsO43-because metabolic pathways intended for PO43- cannot distinguish between the two molecules (7) and arsenate may be incorporated into some early steps in the pathways (6 and refs therein). However, it is thought that downstream metabolic processes are generally not compatible with As-incorporating molecules because of differences in the reactivities of P- and As-compounds (8). These downstream biochemical pathways may require the more chemically stable P-based metabolites; the lifetimes of more easily hydrolyzed As-bearing analogs are thought to be too short. However, given the similarities of As and P, and by analogy with trace element substitutions, we hypothesized that AsO43-could specifically substitute for PO43- in an organism possessing mechanisms to cope with the inherent instability of AsO43-compounds (6).Here, we experimentally tested this hypothesis by using AsO43-, combined with no added PO43-,to select for and isolate a microbe capable of accomplishing this substitution.

砷是磷的化學類似物,在元素週期表上位於磷的正下方。砷不但和磷有相似的原子半徑且都是帶負電性.而在生物體中,大多數的磷是以(PO43-)磷酸鹽的型式存在, 砷鹽(AsO43-)在生物學上相關的pH範圍及氧化還原梯度的表現和磷酸鹽相類似. 在AsO43-的生物毒性上，AsO43-和PO43-兩者間有相似的物理化學貢獻性質.因為在代謝途徑上(PO43-)的設定是無法辨識此這兩者之間的分子,並且AsO43-可能在某些代謝途徑的初始階段進入結合.然而,我們認為在代謝途徑的下游通常不會和含As的分子相容,因為P化合物和As化合物的恢復活性有所差異.生物化學途徑下游可能需要相當穩定的P鍵結代謝物,更易水解的As結合類似物,則認為其生命週期太短. 然而假設砷和磷具相似性,並且從類似物中做微量元素的替換，我們假設AsO43-能具體替代PO43-在生命體中所擁有的機制,以處理AsO43-原有的不穩定性. 在此我們使用了AsO43-進行試驗以確認我們的假設,並且結合不添加PO43-的試驗,去挑選和分離出具有能達成兩者間替換目的的微生物.

Geomicrobiology of GFAJ-1

Mono Lake, located in eastern California is a hypersaline and alkaline water body with high dissolved arsenic concentrations (200 μM on average, 9). We used lake sediments as inocula into an aerobic defined artificial medium at pH 9.8 (10, 11) containing 10 mM glucose, vitamins,trace metals but no added PO43-nor any additional complex organic supplements (e.g. yeast extract, peptone) with a regimen of increasing AsO43-additions initially spanning the range 100 μM to 5 mM. These enrichments were taken through many decimal-dilution transfers greatly reducing any potential carryover of autochthonous phosphorus (11). The background PO43- in the medium was 3.1 (± 0.3) μM on average, with or without added AsO43-, coming from trace impurities in the major salts (11, Table S1).

地質微生物GFAJ-1

Mono湖，位於加利福尼亞東部,具有高鹽度和高濃度的可溶性砷(平均200 μM )的鹼性水體.收集湖中沉積物,接種至好氧的人造合成培養基中(pH 9.8,包含10 mM 葡萄糖，維生素，微量金屬元素,但沒增加PO43-也無另外添加任何複雜的有機補充營養原(例如酵母萃出物，peptone),並且以漸進式持續增加AsO43-濃度的機制進行培養,從最初的100 μM 增到5 mM . 並經由大量的連續稀釋法轉移去除任何潛在殘留的含磷物質. 在有或沒有添加AsO43-的培養基中,背景的PO43-濃度3.1 (± 0.3) μM,來自微量不純的較大鹽類. (11, Table S1).

+/-c

* a Correlation coefficient of sample addition curve was between 0.58 and 0.9. b Sample was from batch grown in June 2010. All other samples were grown in a separate batch in July 2010. C Errors are calculated as the standard deviation on replicate analyses or 10%, whichever is larger.d Error is the expanded error including uncertainties on measurements. “Repeat” indicates a replicate sample digestion and analysis. Sample names in italics designates that the sample was analyzed in several dilutions and on multiple analytical sessions with separate calibration curves, but

not as standard addition curves.

a.樣品增長曲線的相關系數在0.58和0.9之間。

b.樣品來自在2010年6月的生長批次.其他全部的樣品是在2010年7月單離的批次.

c.誤差的預測是取重覆分析的標準偏

差或所測值得的10%(無論最大值為何).d.測量數據上的誤差包括不確定性擴大的

錯誤。“Repeat”指有重覆消化和分析的樣品.樣品的名字以斜體表示代表樣品經由數次稀釋分析和多種分析階段和分隔標準曲線,但沒做標準添加曲線.

The sixth transfer of the 5 mM AsO43-(no added PO43-) condition was closely monitored and demonstrated an approximate growth rate (μ) of 0.1day-1. After 10-7 dilutions, we used the 5 mM AsO43-enrichment to inoculate an agar plate that contained the same chemical composition as the artificial medium. An isolated colony was picked from the agar plates, reintroduced into an artificial liquid medium with no added PO43-where we then progressively increased the AsO43-concentration to determine the optimal level for growth. Currently this isolate, strain GFAJ-1 identified by 16S rRNA sequence phylogeny as a member of the Halomonadaceae family of Gammaproteobacteria (see Fig. S1, 11),

第六次轉養至5 mM AsO43- (無加PO43-)條件下,緊密的監測並證實在0.1天時有相近的生長比率(μ).在經過10-7稀釋倍數後,接種至與人造合成培養基具有相同化合物質的瓊脂培養基(5 mM AsO43-)中.再從plate中挑選單離菌落,轉養至液態培養基中(無添加PO43-),在逐漸的增加AsO43-濃度來決定最適的生長濃度範圍.目前此株分離的細菌strain GFAJ-1,利用16S rRNA做序列分析比對鑑定,在演化樹系統上是屬於Gammaproteobacteria的Halomonadaceae家族,

Figure S1. Phylogenetic analysis of strain GFAJ-1. Maximum likelihood phylogenetic tree based on 16S rRNA genes identified GFAJ-1 as a member of the Halomonadaceae family in the Gammaproteobacteria.

is maintained aerobically with 40 mM AsO43-, 10 mM glucose and no added PO43-(+As/-P condition). Members of this family have been previously shown to accumulate intracellular As(12).

並保存培養在有氧的狀態,含40 mM AsO43-, 10 mM glucose不添加PO43-(+As/-P condition)下.先前的研究指出此家族的細菌會在胞內累積As.

GFAJ-1 grew at an average μmax of 0.53 day-1 under +As/-P, increasing by over 20-fold in cell numbers after six days. It also grew faster and more extensively with the addition of 1.5 mM PO43- (-As/+P, μmax of 0.86 day-1, Fig. 1A, B).

GFAJ-1在有As無P(+As/-P)的條件下,平均最大比生長速率(μmax)為0.53 day-1,在第六天後菌量增加超過20倍.此外在有磷無砷1.5 mM PO43-條件下,細菌則會更快速大量的增長- (-As/+P, μmax of 0.86 day-1, Fig. 1A, B).

Figure 1. Growth, and electron microscopy of strain GFAJ-1. (A, B) Growth curves of GFAJ-1 grown on the defined synthetic medium amended with either 1.5 mM phosphate (solid circles), 40 mM arsenate (solid squares) or neither phosphate nor arsenate (open triangles). Cell growth was monitored both by an increase in (A) optical density and (B) cell numbers of the cultures. Symbols represent the mean ± the standard deviation of n=6 experimental and n=2 controls (A) and n=3 experimental and n=1 control (B). This was a single experiment with six replicates, however material was conserved to extend the duration of the experiment to allow material for cell counting samples. Scanning electron micrographs of strain GFAJ-1 under two conditions discussed in the text. (C) +As/-P and (D) -As/+P. Transmission electron micrography of +As/-P GFAJ-1 (E) showed internal vacuole-like structures. Scale bars are as indicated in the figure (11).

圖1.細菌strain GFAJ-1生長曲線和電子顯微攝影.(A, B)分別是GFAJ-1在含有1.5 mM phosphate (實心圓圈), 40 mM arsenate (實心正方形)或者兩者皆未添加(空心三角形)的培養基中之生長曲線.細菌細胞生長的測定是以兩種方式進行: (A) OD 測細胞密度和(B)細胞計數測定.標示代表標準偏差,(A)實驗組n=6,對照組n=2,(B)實驗組n=3,對照組n=1.每個實驗單獨進行重複六次.而樣品則繼續保存並延長實驗的持續時間,之後進行細菌細胞計數。分別對兩種條件下的strain GFAJ-1菌體進行掃描式電子顯微鏡偵測(C) +As/-P and (D) -As/+P.另外GFAJ-1於有砷無磷的環境下,利用穿透式電子顯微鏡偵測其胞內的囊泡結構.

However, when neither AsO43- nor PO43-was added, no growth was observed (Fig. 1A, B). We include both optical density and direct cell counts to unambiguously demonstrate growth using two independent methods. Cells grown under +As/-P were oblong and approximately two by one microns when imaged by scanning electron microscopy (Fig 1C, 11). When grown under +As/-P conditions, GFAJ-1 cells had more than 1.5-fold greater intracellular volume (vol. ≈ 2.5 ± 0.4 μm3) as compared to -As/+P (vol. ≈1.5 ± 0.5 μm3) (Fig. 1D, 11). Transmission electron microscopy revealed large vacuole-like regions in +As/-P grown cells that may account for this increase in size (Fig. 1E). These experiments demonstrated arsenate-dependent growth, morphological differences in GFAJ-1 driven by AsO43-in the growth medium, and the fact that the level of PO43- impurities in the medium was insufficient to elicit growth in the control (-As/-P).

然而當不添加AsO43-和PO43-時，觀察不到細菌生長(Fig. 1A, B).本研究利用OD和直接記數法來確切的證實細菌的生長情形.當細菌培養在+As/-P時,利用掃描式電子顯微鏡觀察到的菌體呈桿狀約1×2μm大(Fig 1C, 11).當細菌培養在+As/-P下時GFAJ-1的胞內體積(vol. ≈ 2.5 ± 0.4 μm3)比在-As/+P時大1.5倍(vol. ≈1.5 ± 0.5 μm3)

經穿透式電子顯微鏡觀察,在+As/-P條件下生長的細胞有大型的囊泡產生,這跟細胞尺寸變大可能有所關聯. (Fig. 1E).這些實驗證明利用含AsO43-培養基生長的細菌GFAJ-1對As的依賴生長會造成細菌在型態上的差異.而在對照組中(-As/-P),事實上在含有低量PO43-雜質的培養基中並無法誘發細菌的增生.

Cellular stoichiometry and elemental distribution

To determine if GFAJ-1 was taking up AsO43-from the medium, we measured the intracellular As content by ICP-MS (11). In +As/-P grown cells, the mean intracellular As was 0.19 (± 0.25)% by dry weight (Table 1),

細胞組成的化學計量和元素的分佈

為了確定假設細菌GFAJ-1有從培養基中獲取AsO43-,利用ICP-MS測量細菌細胞內的As含量.在+As/-P條件下生長的細胞,乾重細胞胞內As含量佔0.19 (±0.25)% (Table 1),

while the cells contained only 0.02 (±0.01) % P by dry weight. This P was presumably scavenged from trace PO43- impurities in the reagents; and not likely due to carryover given our enrichment and isolation strategy (see above, 11). Moreover, when grown +As/-P this intracellular P is 30-fold less than our measured P values for this microbe when grown -As/+P (see above) and far below the 1-3% P by dry weight required to support growth in a typical heterotrophic bacterium (13). By contrast, GFAJ-1 cells grown under -As/+P conditions had a mean P content of 0.54 (± 0.21) % by dry weight. There was variation in the total As content of the +As/-P cells, possibly a result of collection during stationary phase and losses during the repeated centrifugations and washing cycles due to the potential instability of the cellular

structures given their swollen state (Fig. 2C, E).

同時P的含量則只有0.02 (±0.01) %. P的含量推測可能是從藥劑中所含微量PO43-雜質所得,不像是由於細菌的增生和分離的過程所造成的.此外當細菌生長在+As/-P時,P含量低於生長在-As/+P時的含量約30倍,並且遠低於典型異營微生物生長時所需P的含量(乾重細胞P含量1-3%).相較下, GFAJ-1生長在-As/+P條件下時,乾重細胞內P含量則為0.54 (± 0.21) %.在+As/-P下的細胞其As總含量的變化,可能是在穩定期間As累積所導致,並且在重覆離心和清洗下可能導致細胞組織的不穩定使得他們的構造膨脹。(Fig. 2C, E).

Figure 2. NanoSIMS analyses of GFAJ-1: extracted DNA and whole cells elemental ratio

maps. (A) A garose gel loaded with DNA/RNA extracted from GFAJ-1 grown +As/-P (lane 2) and -As/+P (lane 3) as compared to a DNA standard (Lane 1). Genomic bands were excised as indicated and analysed by NanoSIMS. Ion ratios of 75As-:12C-of excised gel bands are indicated below with 2 sigma error shown (all values multiplied by 10-6). NanoSIMS images of whole GFAJ-1 cells grown either

+As/-P (B, D, F) or -As/+P (C, E, G). The ion ratios of 75As-:12C-(B,C), 31P-:12C-(D,E), and secondary electron, SE (F,G). Ratios in B, C multiplied by 10-4 and D,

E multiplied by 10-3. The color bars indicate measured elemental ratios on a log scale as indicated. Length scale is as indicated on images (11).

GFAJ-1的高解析度二次離子質譜儀分析:DNA萃出物和所有細胞元素的影像分析(A)圖為GFAJ-1DNA萃出物的DNA電泳膠圖，(lane 2)為生長在+As/-P條件下, (lane 3) 生長在-As/+P, (Lane 1)則為DNA標準品.之後挖膠去做NanoSIMS分析. 目標膠體內75As-:12C-的離子比率顯示在下方(所有數值乘上10-6). NanoSIMS 影相分別顯示GFAJ-1生長在+As/-P (B, D, F)和-As/+P (C, E, G)的情形.75As-:12C-的離子比率圖(B,C),31P-:12C-則為(D,E), SE (F,G)則為其電顯圖. B, C圖的比率乘上10-4, D,E則乘上10-3

In contrast, the integrity of the -As/+P cells appeared robust (Fig. 2D) and thus intracellular P measured for these cells likely reflects their content. However, the low total intracellular P in +As/-P cells was consistently far below the quantity needed to support growth, suggesting that these low values are correct despite variation in data from the +As/-P cells. Low intracellular P in concert with high intracellular As was further confirmed by high-resolution secondary ion mass spectrometry and X-ray analyses as discussed below.

相對的,在-As/+P條件下生長的細胞則較為健全完整(Fig. 2D)因此胞內P的測定可能反映出這些細胞的內含物.然而,在+As/-P條件下胞內低含量的總P遠低於維持細菌生長所需的量.這顯示從+As/-P條件下生長的細胞中所得的這些低量值是正確的,不管數據的變化.具有胞內低含量的P和高含量As情況的細菌更進一步的利用high-resolution secondary ion mass spectrometry and X-ray進行分析.分析如下

We used radiolabeled 73AsO43-to obtain more specific information about the intracellular distribution of arsenic (11). We observed intracellular arsenic in protein, metabolite, lipid and nucleic acid cellular fractions (Table 2).

利用放射線同位素73AsO43-來得到更多有關胞內As分部的特殊訊息.得到胞內As 在蛋白質,代謝物,脂質和核酸等細胞組成中的分佈. (Table 2).

細胞清洗後的產物中所有含放射性同位素主要的次組成成份.小分子代謝物包含一些潛在的ATP,NADH,acetyl-CoA and 含其他的arsenylated類似物.

Stationary phase cells incorporated approximately a tenth of the total intracellular

73AsO

43- label into nucleic acids but more than three quarters of the 73AsO

4

3- into the

phenol extracted “protein” fraction, with a small fraction going into lipids. We caution that the large “protein” fraction is probably an overestimate, as this extraction step likely contains numerous small, non-proteinaceous metabolites as well. To determine if this distribution pattern reflected a use of AsO43-in place of PO43-in DNA, we estimated the average sequenced bacterial genome to be 3.8 Mbps, which would contain approximately 7.5 x 106 atoms or 12.5 x 10-18 moles of P. Assuming one complete genome per cell, this would equal 0.39 fg of P in the genome. By ICP-MS, we measured about 9.0 fg P per cell in the -As/+P condition,which implies that only ~ 4 % of total intracellular P is associated with the genome. Since these cells were harvested in stationary phase (11), the fraction of P associated with RNA is likely small (14). Hence, roughly 96 % of P is presumably distributed between the “lipid” and “protein” fractions. If AsO43- is substituting for PO43-in DNA then we can assume that roughly the same fraction of the total intracellular AsO43- would reflect a similar distribution to our estimated PO43-distribution. The distribution of intracellular

73AsO

43- in our experiments was consistent with these estimates. If AsO

4

3-is fulfilling

the biological role of PO43-then AsO43-should act in many analogous biochemical roles including DNA, protein phosphorylation, small molecular weight metabolites (e.g. arsenylated analogs of NADH, ATP, and intermediates like glucose and acetyl-CoA) and phospholipids.

穩定期的細胞合併了大約1/10的73AsO43-標定在核酸上,然而有更多超過3/4的

73AsO

4

3-則進到以phenol(酚)萃取的蛋白質片段上,而有一小部分則進到脂肪中.

本研究謹慎評估這大量的蛋白質片段可能估計過高,在萃取步驟過程中很可能包含了許多小型的非蛋白質性的代謝產物.為了假設並確認這圖樣的分佈反映出了在DNA中AsO43-取代了PO43-的位置.本研究估計算了細菌基因組序列的平均量為3.8 Mbps,這樣的量大約包含了7.5 x 106個原子量或12.5 x 10-18莫耳的P.每一個完整的細胞中其基因組所含的P平均值為0.39 fg( 0.39×10-15g),經由ICP-MS分析得到在-As/+P條件下,每個細胞含有9.0 fg P,這暗示著在細胞內和基因組聯結的P含量只占總P含量的4%.在生長穩定期細胞中的RNA和P聯結的部分可能很小.因此大約有96%的P可能分散在蛋白質和脂質片段之間.假設在DNA中AsO43-是替代PO43-的位置,因此可以初步假設細胞內在相同部位的總AsO43-分佈,反映出和我們評估的PO43-有相似的分佈.在本實驗中細胞內73AsO43-的分佈和之前的實驗評估一致.假設AsO43-在生物學上履行了PO43-的角色, 那麼AsO43-應該會扮演很多類似的生物化學角色包括在DNA，蛋白質磷酸化，小分子代謝物(例如：NADH，ATP的arsenylated類似物及葡萄糖和acetyl-CoA的媒介物) 以及磷脂上。

Our data suggested that arsenic was present in a number of biomolecules and in

particular we sought to confirm the presence of arsenic in the DNA fraction. Initially, we measured traces of As by ICP-MS analysis of extracted nucleic acid and protein/metabolite fractions from +As/-P grown cells (11, Table S1). We then used high-resolution secondary ion mass spectrometry (NanoSIMS) to positively identify As in extracted, gel purified genomic DNA (Fig. 2A). These data showed that DNA from +As/-P cells had elevated As and low P relative to DNA from the -As/+P cells. NanoSIMS analysis of the DNA showed that the As:P ratio on an atom per atom basis was significantly higher in the +As/-P versus -As/+P grown cells (Fig. 2A, 11?Table S2).

本研究結果顯示砷在許多生物分子中都存在,此外我們特別去確認DNA部分砷的存在.起初,利用ICP-MS分析培養在+As/-P條件下的細胞萃取物：核酸、蛋白質和代謝物等部分As的蹤跡.(11, Table S1).之後利用高分辨二次離子質譜儀high-resolution secondary ion mass spectrometry (NanoSIMS)去確認從DNA genomic膠體中純化出的萃出物內所含的As. (Fig. 2A).數據顯示在+As/-P條件下的細胞DNA相較於-As/+P條件下的DNA有較高的As和較低的P含量.經由NanoSIMS分析的DNA所顯示的砷磷原子比率(As:P),結果暗示在+As/-P對-As/+P 條件下生長的細胞有較高的比率.

?Measurements are mea n values for splits of excised agarose gel bands loaded with purified GJAJ-1DNA and RNA; the other portion was run by ICP-MS. a, c Elemental concentrations are for hydrated agarose. B All error values reported in this table are 2 * standard error. D Blank values were measured on agarose gel segments excised

from outside of electrophoresis lanes; these are an upper limit of expected background ion concentrations caused by trace impurities in agarose, loading dye or ethidium bromide stain.

Whether expressed as an ion ratio relative to C, (75As-:12C-, Fig. 2A) or 31P-:12C- (11, Table S2) or normalized by relative ion yield and expressed as a concentration in parts per billion (11, Table S2), we saw a similarly consistent trend, with significantly higher As in the +As/-P DNA, and higher P in the -As/+P DNA. In both cases, the non-amended element concentration was equal or less than background levels. These measurements therefore specifically demonstrated that the purified DNA extracted from +As/-P cells contained As. Our NanoSIMS analyses, combined with the evidence for intracellular arsenic by ICP-MS and our radiolabeled

73AsO

43-experiments demonstrated that intracellular AsO

4

3- was incorporated into key

biomolecules, specifically DNA.

並且表示相對於C原子的比率,(75As-:12C-, Fig. 2A) 或31P-:12C- (11, Table S2)或者標準化相對離子產率表示每10億分之幾的濃度(11, Table S2),研究發現一個相似且始終如一的趨勢,顯示在+As/-P條件下DNA有較高的As含量,在-As/+P條件下DNA則有較高的P含量.在此兩種情況下,無修正元素的濃度是等於或低於背景值的. 因此這些測量數據都明確的證明在+As/-P條件下生長的細胞,純化後的DNA 萃出物含As. 本研究的NanoSIMS分析,結合了ICP-MS來證實胞內砷的蹤跡和放射性同位素73AsO43-的實驗證實胞內AsO43-是構成一特殊DNA的關鍵生物分子.

Characterization of the intracellular arsenic chemical environment

We next used synchrotron X-ray studies to determine the speciation and chemical environment of the intracellular arsenic (11). Micro X-ray absorption near edge spectroscopy (μXANES) of +As/-P grown cells exhibited an absorption edge characteristic of As(V) coordination with no evidence of As(III) observed. Best fits of the micro extended X-ray absorption fine structure (μEXAFS) spectra are listed in Table3 and shown in Figure 3.

胞內砷的化學環境特性

接著使用同步加速器X射線來確定細胞內砷的型態分析和化學環境的研究( 11 ) .利用近緣X光吸收細微結構光譜(μXANES)分析在+As/-P下生長的細胞,結果顯示有As(V)配位的近緣吸收特性,沒有As(III)的吸收跡象.最佳的近緣X光吸收細微結構光譜如表3和圖3所列.

表3: GFAJ-1胞內砷的近緣X光吸收細微結構:

細節:表S02=1,全區性振幅因子和E0=13.97,標定的距離.Type 配位型態, Number 配位數.R,原子間的距離.σ2,原子外圍的靜電亂數測量值(德拜瓦倫因子(Debye-Waller factor)，為第i層原子在距離上的均方根偏差(Root-meansquare)，它是由於原子間結構上亂序(Disorder)(屬靜態的)及熱震動(屬動態的)，所產生的結果).見表S2比對含P的生物分子中的P.(11)

Figure 3. X-ray analysis of GFAJ-1 +As/-P described similarity of As coordinated like P in DNA. (A) EXAFS comparisons of the Fourier transformed data for two model compounds, As-S and As-Fe, whole GFAJ-1 cells (washed and fixed) and a fit of DNA with arsenic replacing phosphorus, in silico. Identification of each spectrum is indicated on the figure and from top to bottom are As-S, As-Fe, GFAJ-1 data (collected on whole cells) and fit to the GFAJ-1 data (in red). (B) XRF maps indicated the correlation between arsenic (As), iron (Fe) and zinc (Zn) and

not with phosphorus (P) with some variability but consistent with the trend that these elements are often found together (See Figure S3 in the SOM for element correlation plots). The length scale bar in the “Zn” quadrant, of the maps is as designated and applies to all parts of the figure. Given the spatial resolution of these images, the structures identified as containing high As, Fe, and Zn are aggregates of cells. Ranges as indicated in the color bar run from cold to hot, in units of μg cm-2, as follows: As, 0 to 1.6; P, 0 to 40; Fe, 0 to 32.1, and Zn, 0 to 2.8. Standards were used to calibrate signal and background (11).

圖3經由X射線分析GFAJ-1在+As/-P條件下,顯示As在DNA上的配位類似於P (A)經由EXAFS的傅利葉轉換數據在電腦模擬運算中,比較兩種化合物模型: As-S and As-Fe, 所有GFAJ-1細胞(含清洗和有雜質的部份)和DNA中As取代P的部位

的匹配性.確認每個光譜波形的標示,上部到底部波峰分別是As-S, As-Fe的波峰位置, GFAJ-1 data(收集所有細胞)和GFAJ-1 data 配位的部份(紅色所示DNA中As 取代P的配位)(B) XRF圖譜指出arsenic (As), iron (Fe) and zinc (Zn)間的關連性,沒有偵測到P和一些變異,但這些元素常常被一起發現並有一致性的趨勢(見圖S3 在SOM分析中元素間關聯性的圖).在4個象限圖中鋅的部份有一尺標刻度,這一尺標也套用在所有其他圖的刻度.由這些圖中可得到一空間解析,由空間結構可鑑定在這些由細胞收集得到的物質含有高量的As, Fe, and Zn.條帶的顏色範圍指示由冷色系到艷色系為代表,單位為μg cm-2,依序為As:0 ~1.6; P: 0 ~ 40; Fe: 0 ~ 32.1, 和Zn: 0 ~ 2.8.利用標準品來校正訊號和背景值(11)

The first neighbor shell around the arsenic in +As/-P cells consisted of four oxygen ligands (Table 3), but has a second shell that is inconsistent with our As-Fe and As-S models, free arsenate ions or published spectra for organo-arsenicals (Fig. 3A, 15, 16). While other arsenical compounds, such as dimethylarsinate (DMA) also have As-O and As-C bonds, they have edge positions which are shifted to lower energy from the observed As(V) and have much shorter observed As-C bond distances (16). In contrast to the models, these As-O and As-C distances are consistent with that reported from the solved crystal structure of DNA for the analogous structural position of P relative to O and C atoms (Fig. 3A, 16, 17). Therefore, our X-ray data support the position of arsenate in a similar configuration to phosphate in a DNA backbone or potentially other biomolecules as well. These data also indicated evidence for the presence of arsenate in small molecular weight metabolites(e.g., arsenylated analogs of NADH, ATP, glucose, acetyl-CoA) as well as arsenylated proteins where arsenate would substitute for phosphate at serine, tyrosine and threonine residues (1, 11, Table S3).

在+As/-P條件下生長的細胞中,經由有機砷發散的光譜或游離砷離子中分析得到第一層鄰近圍繞在As周圍的是由4個氧所組成的配位體(Table 3),然而第二層則和

十大词性 句子成分及基本句型讲解及练习(附答案)..

十大词性、句子成分分析、五大基本句型及句子类型讲解与练习 一、新课导入 个体名词 集体名词 普通名词 物质名词 1.名词 抽象名词 专有名词 功能：表示人或事物的名称 人称代词： 主格 宾格 （注意it 的用法） 物主代词： 形容词性物主代词 名词性物主代词 指示代词：this, that, these, those 单数：myself`, yourself, himself, herself, itself, oneself 反身代词： 复数：ourselves, yourselves, themselves 2.代词 疑问代词：who, whom, whose, what, which. 简单不定代词：One/ones, Either/neither, both/all, each/ every another, other, the other, others, the others 复合不定代词： somebody, anybody, nobody 不定代词 someone, anyone, no one something, anything, nothing everybody, everyone, everything 功能：用来代替名词、形容词或数词等 3.数词： 基数词 序数词 功能：表数量或顺序 4.冠词： 定冠词 不定冠词 零冠词 //功能：用在名词前，帮助说明名词所指的人或事物 表地点：at/ in/ on/ to, above/ over/ on, below/ under/ underneath …. 表时间： in/ on/ at, in/ after, from/since from, after/ behind, 5.介词： 表运动： across/ through/ over/ pass … 表方位：…. 其他 功能：用在名词、代词等前面，说明句子中词与词之间的关系 可数名词 不可数名词

五种基本句型和句子成分

句子成分 英语句子成分有主语，谓语，宾语，宾语补足语，表语，定语，状语等。 顺序一般是主语，谓语，宾语，宾语补足语，而表语，定语，状语的位置要根据情况而定。 1、主语：表示句子主要说明的人或事物，一般位于句首。但在there be结构、疑问句（当主语不疑问词时）和倒装句中，主语位于谓语、助动词或情态动词后面。主语可由名词、代词、数词、不定式、动名词、名词化的形容词和主语从句等表示。例如： Country music has become more and more popular.（名词） We often speak English in class.（代词） One-third of the students in this class are girls.（数词） To swim in the river is a great pleasure.（不定式） Smoking does harm to the health.（动名词） The rich should help the poor.（名词化的形容词） When we are going to have an English test has not been decided.（主语从句） It is necessary to master a foreign language.（it作形式主语，真正的主语为后面的不定式） 2、谓语：谓语说明主语的动作，状态或特征。可以有不同的时态，语态和语气。We study for the people. I can speak a little English. We are reading books. He has gone to Beijing. 3、表语: 它位于系动词(比如be)之后，说明主语身份、特征、属性或状态。 My sister is a nurse. （名词） Is it yours?（代词） The weather has turned cold.（形容词） The speech is exciting.（分词） Three times seven is twenty one?（数词） His job is to teach English.（不定式） His hobby is playing football.（动名词）

(完整版)英语句子成分和英语句子结构讲解和练习试题

英语句子成分和英语句子结构讲解及练习 简单句的五个基本句型 ?主语＋不及物动词She came．. ?主语＋及物动词＋宾语She likes English. ?主语＋系动词＋主语补语 ?She is happy． ?主语＋动词＋间接宾语＋直接宾语 ?She gave John a book． ?She bought a book for me. ?主语＋动词＋宾语＋宾语补语 ?She makes her mother angry． ?The teacher asked me to read the passage. ?（There +be There lies a book on the desk. 主谓宾 ?名／代－－动词－－名／代 ?we-- saw --you. ?we-- did --the work. ?主系表 ?名／代－系动词－形容次／名词／代词 ?you are beautiful ?you seems worried. ?you are a stufent.

?相同点都三部分，主语也一样．不同动词和系动词，时态一样，否定式不同．动词加助动词，系动词不用．表语可以是形容词，宾语不行．只有宾语有补足语 2、句子成分：英语句子成分分为七种： ?主语、谓语、宾语、定语、状语、表语、宾语补足语。 ?1、主语是句子所要说的人或事物,回答是“谁”或者“什么”。通常用名词或代词担任。 ?如：I’m Miss Green.(我是格林小姐) ?2、谓语动词说明主语的动作或状态,回答“做(什么)”。主要由动词担任。 ?如：Jack cleans the room every day. (杰克每天打扫房间) ?3、表语在系动词之后,说明主语的身份或特征,回答是“什么”或者“怎么样”。通常由名词、代词或形容词担任。 ?如：My name is Ping ping .(我的名字叫萍萍) 4、宾语表示及物动词的对象或结果,回答做的是“什么”。通常由名词或代词担任。 ?如：He can spell the word.(他能拼这个词) ?有些及物动词带有两个宾语,一个指物,一个指人。指物的叫直接宾语,指人的叫间接宾语。间接宾语一般放在直接宾语的前面。 ?如：He wrote me a letter . (他给我写了一封信) ?有时可把介词to或for加在间接宾语前构成短语,放在直接宾语后面,来强调间接宾语。如：He wrote a letter to me . (他给我写了一封信) 5、定语修饰名词或代词,通常由形容词、代词、数词等担任。如： Shanghai is a big city .(上海是个大城市) 6、状语用来修饰动词、形容词、副词,通常由副词担任。 如：He works hard .(他工作努力)

(word完整版)It+willbe+时间段+before等表示“在……之后……才”的句型总结,推荐文档

It + will be + 时间段 + before等表示“在……之后……才”的句型总结 一、用于句型“It + will be + 时间段 + before...”句型中，表示“要过多久才…”，也可用于“It + may be + 时间段 + before...”，表示“也许要过多久才……”。Before 后的句子中用一般现在时态。 其否定形式“It will/would not be +时间段+ before…”表示“不久就……，过不了多久就……”。 【典型考例】 （1）The field research will take Joan and Paul about five months; it will be a long time _____ we meet them again.（2007安徽卷） A. after B. before C. since D. when （2）—How long do you think it will be ______China sends a manned spaceship to the moon? （2006福建卷） —Perhaps two or three years. A. when B. until C. that D. before （3）It ________ long before we _______ the result of the experiment.( 上海春招2002) A. will not be...will know B. is...will know C. will not be...know D. is...know (4) Scientists say it may be five or six years_________ it is possible to test this medicine on human patients. （2004福建） A. since B. after C. before D. when 解析：答案为BDCC。考题 （1）(2)before 用于肯定的“It + will be + 时间段 + before...”句型中，表示“要过多久…才…”。 （3）before在本题中用于否定句,意为“过不了多久就会……”, 状语从句要用一般现在时代替一般将来时，可知C项为正确答案,句意是:要不了多久我们就会知道试验的结果了。 （4）宾语从句中含有句型“It + may be + 时间段 + before...”，表示“也许要过多久才……”，故选择答案C。 二、用于句型“it was +时间段+ before …”表示“过了（多长时间）才……”。其否定形式“ it was not +时间段+ before …”意为“不久就……”, “没过（多长时间）就……”。 【典型考例】 It was some time ___________we realized the truth. （2005山东） A. when B. until C. since D. before 解析：答案为D。before用于句型“it was +时间段+ before …”表示“过了（多长时间）才……”。该题题意是“过了一段时间我们才意识到事情的真相”。故正确答案为C项。

五种基本句型及句子成分分析

五种基本句型及句子成分的分析五种基本句型的分析能力的培养是分析一切复杂句子的基础，是分析千千万万个句子的基础，因为众多的 句子都是由五种基本句型变换过来的。 所谓五种基本句型分析能力就是：看到一个简单句就能立刻分析出这个句子属于五种句型中的哪种；能够立刻分析出句子的主干是什么，能够快速分割，快速整合。 怎样才能具备这种能力呢？ 先理解理论（怎么去做的方法）后多实践做练习，实践是最重要的，能力在大量的实践练习中应该可以掌握。 在培养这个能力过程中还需要记忆任务。如：哪些动词后面要跟动词不定式,哪些动词后面要跟doing ,哪些动词后面需要跟双宾语等等这些都需要记忆，这项记忆任务完全可以通过练习实践来掌握，不做练习，只去死记那些东西不容易掌握。 如果对汉语句子成分分析有一定的了解和掌握，有助于英语句子成分的学习，因为本来汉语句子成分分析那 一套就是从英语那里搬过来的。 五种基本句型 1，主（语）＋系（动词）＋表（语） 首先我们来认识主语，主语是五种基本句型都具备的。从位置上来看，主语一般在句子开头，也就是一个句子的“头把交椅”：从意义上理解，主语一般是一个句子的“陈述对象”，后面的一切都是针对主语来说的，那么常坐这把“交椅”的会是什么词呢，一般是名词或代词。 系动词是联系动词的简称，顾名思义，它是起到联系作用的动词，是连接主语和表语的。系动词的数量是有限的，常见的只有十来个，我们把它们分成四组，这样有助于我们记忆系动词： 一、be动词类：am、is 、are 、was 、were 二、表示变化类：become、get 、turn 、grow 、go 三、感官动词类: 眼（look ）耳(sound)鼻(smell)舌(taste)身(feel) 四、表示延续性的动词：remain 、stay 、keep 表语有两个特征：一从位置上来讲，它总是在系动词后面（系动词和表语关系甚为紧密，没有系动词，就没有表语：没有表语，就没有系动词，正所谓“皮之不存，毛将焉附”）。二从意义上讲，表语通常是来说明主语的性质、特征的。表语通常是由形容词、介词短语、名词、代词、动词不定式、doing 等来充当。 上面就是对主系表结构的简要介绍，其中，系动词是主系表结构的核心，它是这个结构的标志，看到了系动词，也就能立刻判断出句子的结构是主系表，因此，记住系动词非常重要。 2，主（语）＋谓（语） 谓语，谓语从意义上说有表示动作和状态两种情况；表示状态的就用主语＋连系动词＋表语句型来表示。一般情况下，谓语位于主语之后，从意义上理解，往往具有动作意味（谓语实际有表示动作和状态两种情况；表示状态的就用主语＋连系动词＋表语句型来表）是主语发出的动作。能做谓语的在十大词类中只有一种词类：动词，也就是说只用动词具有作谓语的“专利”，所以我们往往把“谓语”和“动词”连在一起叫做“谓语动词”。虽然主语是“头把交椅”，但是谓语是一个句子中最重要的成分，一个句子可以缺少主语，但是谓语是不能少的。 为了更好地理解这个句型以及下面的三个句型，在这里我需要补充说明两点：

before用法归纳

before用法知多少？ 在高考中，状语从句是每年高考单项填空部分必考的题目之一，考查的重点是考生容易混淆并且近似的连词在逻辑行文和语篇结构中的使用。before作连词的用法一直是高考的重点，也是学生感觉掌握起来比较头疼的地方。下面选取近几年各省市的高考试题进行归纳分析，使考生通过典型实例，把握高考对before所引导的句型的命题规律，帮助同学们更好地解答此类题目。 1. before作为连词时的基本意义是“在……之前”，用于表示时间或顺序。 You can’t borrow books from the school library ______ you get your student card. (2009上海，32) A. before B. if C. while D. as 【解析】选A。考查连词，该句的意思是：在你得到你的学生卡之前你不能从学校图书馆借书。before表示先后顺序。 2. 表示“过了多久才……”，说明主句的持续时间比较长而从句的动作缓缓来迟。 (1) The American Civil War lasted four years _______ the North won in the end. (2005广东，30) A. after B. before C. when D. then 【解析】选B。本题考查连词before表示“在多久之后才……”的用法，根据本句含义“美国南北战争持续了四年，北方才最终取得胜利”，可知本题应选B。 (2) Several weeks had gone by I realized the painting was missing. (2004宁夏，39) A. as B. before C. since D. when 【解析】选B。before表示“过多久才……”。句意：几个星期已经过去了，我才意识到油画丢了。内含的意思是油画丢了好几个星期了，我才意识到。 3. 表示从句动作还没来得及发生或完成，主句动作就已经发生或完成了，意为“尚未……就”，“没来得及……就”，常用于before sb. can/ could…。 —Why didn’t you tell him about the meeting? ( 2006四川，35) — He rushed out of the room _________ I could say a word. A. before B. until C. when D. after 【解析】选A。本题考查连词before表示“还没来得及……就……”的用法。句意为：我还没来得及说一句话，他就冲出了房间。 4. 表示“以免，以防，趁……还没有……”，强调动作的必要性，以避免或防止从句动作的发生。 He made a mistake, but then he corrected the situation _____ it got worse.（2003北京） A. until B. when C. before D. as 【解析】选C。由made a mistake和转折词but可知本题句意是“他犯了一个错误，但在事情进一步恶化之前他改变了形势。”故答案正确答案为C项。

but,than引导定语从句和before,until用法辨析

but和than引导定语从句的用法 一、but可被看作关系代词，引导定语从句，在句中作主语，在意义上相当于 who not或that not，即用在否定词或具有否定意义的词后，构成双重否定。 如：①There is no mother but loves her children．没有不爱自己孩子的母亲。 ②There was no one present but knew the story already．在场的人都知道这个故事。 二、than作关系代词时，一般用在形式为比较级的复合句中，其结构为形容词比较级（more）．．．than＋从句，than在从句中作主语，相当于that，代表它前面的先行词。（这时，它兼有连词和代词的性质，也有学者认为这种用法的than是连词，后面省略了主语what。） 如：①The indoor swimming pool seems to be a great deal more luxious than is necessary．室内游泳池过于豪华。 ②He got more money than was wanted．他得到了更多的钱。运用上述知识翻译下列句子： 1．任何人都喜欢被赞扬。（but） 2．我们大家都想去桂林。（but） 3．没有人不同情那些嗷嗷待哺的孩子。（but） 4．我们班上没有一个人不想帮你。（but） 5．无论多么荒凉，多么难以行走的地方，人们也能把它变成战畅（but）6．这件事情比想象的要复杂。（than） 7．这个广告的效果比预想的要好。（than） 8．这个问题看起来容易，实际上很难。（than） 9．他爸妈给他的零用钱总是超过他的需要。（than） 10．因为这项工程非常困难，所以需要投入更多的劳动力。（than） 答案： 1．There is no one but likes to be praised． 2．There is no one of us but wishes to visit Guilin． 3．There is no man but feels pity for those starving children．4．There is no one in our class but wants to help you． 5．There is no country so wild and difficult but will be made a theatre of war． 6．This matter is more complex than is imagined． 7．This advertisement is more affective than is expected． 8．The problem may be more difficult in nature than would appear．9．He got more pocket money from his parents than was demanded．

英语句子成分和句型结构

一、英语句子成分和英语句子结构讲解： （一）句子成分 1.主语（subject）: 句子说明的人或事物。 主语可以由名词、代词、数词、不定式、动名词、分词、主语从句和短语等来担任。 The sun rises in the east.（名词） He likes dancing. （代词） Twenty years is a short time in history. (数词) Seeing is believing. （动名词） To see is to believe. （不定式） What he needs is a book. （主语从句） It is very clear that the elephant is round and tall like a tree. （It形式主语，主语从句是真正主语） 找出下列句中的主语： Jane is good at playing the piano.（名词） She went out in a hurry.（代词） Four plus four is eight.（数词） To see is to believe.（不定式） Smoking is bad for health.（动名词） The young should respect the old.（名词化的形容词） What he has said is true. （句子） 2.谓语（predicate）: 说明主语的动作、状态和特征。 简单谓语:由动词或动词词组组成 I saw the flag on the top of the hill He looked after two orphans. 复合谓语：由情态动词或助动词+动词; He can speak English well. She doesn’t seem to like dancing. 找出下列句中的谓语（注：只有动词才可作谓语。）： 1. We love China. 2. We have finished reading this book. 3. He can speak English. 4. She seems tired. 3.表语（predicative）: 系动词之后的成分，表示主语的性质、状态和特征。 He is a teacher. （名词） Seventy-four! You don’t look it. （代词） Five and five is ten. （数词） He is asleep. （形容词） His father is in. （副词） The picture is on the wall. ( 介词短语) My watch is gone / missing / lost. （形容词化的分词） To wear a flower is to say “I’m poor, I can’t buy a ring. （不定式） The question is whether they will come. （表语从句） ★（常见的系动词有: be, sound（听起来）, look（看起来）, feel(摸起来，

初中句子成分和基本句型完整版

初中句子成分和基本句 型 Document serial number【NL89WT-NY98YT-NC8CB-NNUUT-NUT108】

句子成分和基本句型 句子包含哪些成分？ 现代汉语里一般的句子成分有六种，即主语、谓语、宾语、定语、状语和补语。英语的基本成分有八种：主语（subject）、谓语（predicate）、表语（predicative）、宾语（object）、定语（attribute）、状语(adverbial)、{补语（complement）、同位语（apposite）} 1.主语：主语是在句子中说明全句中心主题的部分．一般由名词，代词，不定式，动名词或从句充当．它的位置一般在句首． Thegirlispretty.Readingisuseful. Toseeistobelieve.Theyaregoodfriends. 2.谓语说明主语做什么，以什么状态或特征存在，由简单动词或者动词短语构成． Theyareteachers.Shelookswell. Hestudieshard.Helaughedathisclassmates. 3.表语说明主语是什么，或者怎么样，由名词，形容词，副词，介词短语，不定式，动名词，短语，或者从句充当．表语位于系动词后面Iamateacher.Shefeelshappy. Everybodyishere.Theyareathomenow. Hewentmad.Seeingisbelieving. 4.宾语是动作，行为的对象，由名词，代词，短语，不定式，动名词，从句充当．它和及物动词一起说明主语做了什么，在谓语之后．Sheplaysthepiano.Heoftenhelpsme. IlikewatchingTV.Shelikestogotoshopthisafternoon. Ithinkthatheisgoodboy. 5.宾语补足语：在英语中，有些句子只有宾语并不能表达完整的意思，还必须在宾语后面家上宾语补足语才能表达完整的意思．它起补充说明宾语干什么，怎么样的作用． 它可以由名词，形容词，副词，介词短语，不定式，现在分词和过去分词充当． Ifyouletmego,I’llmakeyouking.Leavethedooropen. WefoundJohnoutwhenwearrived.Makeyourselfathome. Iheardmynamecalled. 6.状语用来修饰动词，形容词或副词．它表示行为发生的时间，地点，目的，方式，程度．一般由副词，介词短语，不定式或相当于副词的词或短语充当，也可由从句来充当． Hediditcarefully.Hestudiedveryhard. IwasbornonJune9th1982inGuangxi.Heiswritingwithapen. Iwillstayathomeifitrainstomorrow.

before句型辨析与解析

before句型辨析与解析 It + will be + 时间段+ before等表示“在……之后……才”的句型总结 一、用于句型“It + will be + 时间段+ before...”句型中，表示“要过多久才…”，也可用于“It + may be + 时间段+ before...”，表示“也许要过多久才……”。Before 后的句子中用一般现在时态。 其否定形式“It will/would not be +时间段+ before…”表示“不久就……，过不了多久就……”。 （1）The field research will take Joan and Paul about five months; it will be a long time _____ we meet them again. A. after B. before C. since D. when （2）—How long do you think it will be ______China sends a manned spaceship to the moon? —Perhaps two or three years. A. when B. until C. that D. before （3）It ________ long before we _______ the result of the experiment. A. will not be...will know B. is...will know C. will not be...know D. is...know (4) Scientists say it may be five or six years_________ it is possible to test this medicine on human patients. A. since B. after C. before D. when 解析：答案为BDCC。考题 （1）(2)before 用于肯定的“It + will be + 时间段+ before...”句型中，表示“要过多久…才…”。 （3）before在本题中用于否定句,意为“过不了多久就会……”, 状语从句要用一般现在时代替一般将来时，可知C项为正确答案,句意是:要不了多久我们就会知道试验的结果了。 （4）宾语从句中含有句型“It + may be + 时间段+ before...”，表示“也许要过多久才……”，故选择答案C。 二、用于句型“it was +时间段+ before …” 表示“过了（多长时间）才……”。其否定形式“ it was not +时间段+ before …” 意为“不久就……”, “没过（多长时间）就……”。 It was some time ___________we realized the truth. A. when B. until C. since D. before 解析：答案为D。before用于句型“it was +时间段+ before …” 表示“过了（多长时间）才……”。该题题意是“过了一段时间我们才意识到事情的真相”。故正确答案为C项。 表示“在……之后……才”。副词“才”在汉语中强调某事发生得晚或慢。如果在含有before从句的复合句中，强调从句动作发生得晚或慢时，就可以应用这种译法。 The American Civil War lasted four years _______ the North won in the end. A. after B. before C. when D. then 解析：答案为B。本题考查连词before表示“在……之后才……”之的用法，根据本句含义“美国南北战争持续了四年，北方才最终取得胜利”，可知本题应选B。 三、表示“……还没来得及……就……”。目的在于强调从句动作发生之前，主句动作已发生。 —Why didn't you tell him about the meeting? — He rushed out of the room _________ I could say a word. A. before B. until C. when D. after 解析：答案为A。本题考查连词before表示“……还没来得及……就……”的用法。句意是“我还没来得及说一句话，他就冲出了房间”。 四、表示“在……之前就……”。这时主句与before从句中的两个动作按时间先后依次发生。 It was evening______ we reached the little town of Winchester. A. that B. until C. since D. before 解析：答案为D。本题考查连词before表示“在……之前就……”的用法。句意是“我们到达小镇Winchester 之前就已经是傍晚时分了”。 五、表示“趁……”，“等到……”，或“没等……就…… ”等。

句子成分和基本句型

句子成分和基本句型 什么是句子？ 句子是由词按照一定的语法结构组成的，是能表达一个完整概念的语言单位．句子的第一个字母必须大写，结尾要有．？！ I am a teacher. Are you a student? How beautiful the girl is ! *句子成分由词或词组充当 英语的基本成分有七种： 主语(subject)、谓语(predicate)、表语(predicative)、宾语(object)、定语(attribute)、状语(adverbial)补语(complement) １．主语：主语是在句子中说明全句中心主题的部分．一般由名词，代词，不定式，动名词或从句充当．它的位置一般在句首． The girl is pretty. Reading is useful. To see is to believe. They are good friends. 谓语： 谓语说明主语做什么，是什么，怎么样，由简单动词或者动词短语构成． They are teachers. She looks well. He studies hard. He laughed at his classmates. He can speak English. 表语： 表语说明主语是什么，或者怎么样，由名词，形容词，副词，介词短语，不定式，动名词，短语，或者从句充当．表语位于系动词后面． I am a teacher. She is happy. Everybody is here. They are at home now. My job is to teach English. Seeing is believing. 宾语 宾语是动作，行为的对象，由名词，代词，短语，不定式，动名词，从句充当．它和及物动词一起说明主语做了什么，在谓语之后． She plays the piano. He often helps me.

英语句子成分及基本句型

英语句子成分及基本句型 句子成分 A. 主语（Subject）主语是一句话的中心。除祈使句外，主语是句子中不可缺少的成分。主语表示句子中所说的是“谁”或 “什么”。 Jane is good at playing the piano. She went out in a hurry. Thirteen is regarded as an unlucky number by some people. To see is to believe. Smoking is bad for health. The young should respect the old. What he has said is true. B. 谓语（Predicate）谓语表示主语的动作或状态，分为简单谓语与复合谓语两种。简单谓语由动词或动词词组组成；复合 谓语由情态动词或助动词+动词。 I saw the flag on the top of the hill? He looked after two orphans. He can speak English well. C. 表语（Predicative）在系动词后的部分就是表语,说明主语的状态等。 常见的系动词有be(am,is,are,were,was),appear,look,seem,feel,smell,taste,sound,keep,turn,become, get, grow, come, go etc.系动词不用于被动语态.The apple tastes sweet. The war was over. They seem to know the truth. Time is precious. I’m not quite myself today. Who was the first? He is out of condition. The book is what I need. D. 宾语（Object）宾语是动作行为的对象，说明主语“做什么”。宾语分为直接宾语（Direct Object）和间接宾语(Indirect Object)。直接宾语常指物,间接宾语常指人。Give him a cup of coffee. E.宾语补足语（Object Complement）是用在宾语之后补充宾语情况的成分。 I found the book interesting. Do you smell something burning? He made himself known to them. She asked me to lend her a hand. Please make yourself at home. Please keep the dog out. We must keep it a secret. F．定语（Attribute）定语修饰名词或代词。 He is a clever boy. His father works in a steel work. There are 54students in our class. Do you known Betty’ｓsister? There is a sleeping baby in bed. His spoken language is good. I met a friend on my way home. G．状语（Adverbial）用于修饰形容词,副词,动词,介词短语或句子.可表示时间,地点,原因,结果,目的,方式,伴随,条件,程度,让步,频率等。 I will be back in a while. They are playing on the playground. He was late because he got up late. He got up so late that I missed the train. I waited to see you. He often went to school by bus. His parents died, leaving him an orphan. Please call me if it is necessary. This book is very interesting. He went to school in spite of his illness. He always comes late to school. 讲出划线部分的句子成分： 1．Her suggestion sounds quite reasonable. 2．I have a lot to say on this matter. 3．Please give me some advice. 4．She is quite all right now. 5． A lonely person is always alone. 6． A fat, sandy-haired boy in jeans came to see Miss White.

Before用作连词时

Before用作连词时，意思是“在……之前”。(注：此为核心概念，其它皆是繁衍派生之义)其实，它引导状语从句时，在不同的句式中以及汉英表达习惯的不同，before含有不同的含义和用法。（注：翻译意思不同而已）。（注：当主从句动作有先有后的时候，用before，after居多，但其他的呢？居多是多少？有什么使用条件？） （最容易错的就是和when难以区分，这主要是受到中文翻译法的影响，但如何辨别？） 从历年的高考试题可以看出，before是高考考查的热点词汇之一。为了帮助大家掌握，现就对它的用法归纳如下： 一、表示“在……之后……才”。副词“才”在汉语中强调某事发生得晚或慢。如果在含有before从句的复合句中，强调从句动作发生得晚或慢时，就可以应用这种译法。 【典型考例】 The American Civil War lasted four years _______ the North won in the end.(2005广东) A. after B. before C. when D. then 解析：答案为B。本题考查连词before表示“在……之后才……”之的用法，根据本句含义“美国南北战争持续了四年，北方才最终取得胜利”，可知本题应选B。 （注：这种题目比较难，因为学生如果用中文翻译成“当”when的时候，句子也读得顺，如果来明显区分？） 二、表示“……还没来得及……就……”。目的在于强调从句动作发生之前，主句动作已发生。 【典型考例】 —Why didn't you tell him about the meeting? ( 2006四川卷) — He rushed out of the room _________ I could say a word. A. before B. until C. when D. after 解析：答案为A。本题考查连词before表示“……还没来得及……就……”的用法。句意是“我还没来得及说一句话，他就冲出了房间”。 三、表示“在……之前就……”。这时主句与before从句中的两个动作按时间

初中英语句子成分和五种基本结构

句子的成分 1 句子的成分 组成句子的各个部分叫做句子的成分。句子成分包括：主语、谓语、表语、宾语（直接宾语、间接宾语）、宾语补足语、定语和状语。主语和谓语是句子主体部分（在英文中一般的句子必须有主语和谓语）。表语、宾语和宾语补足语是谓语里的组成部分。其他成分如定语和状语是句子的次要部分。下面我们分别讲述一下句子的各个成分： 1 主语 主语是谓语讲述的对象，表示所说的“是什么”或“是谁”。一般由名词、代词、不定式或相当于名词的词或短语来充当。它在句首。 We study in Middle School.（讲述“谁”～） The classroom is very clean. （讲述“什么”很干净） Three were absent.（数词作主语） To teach them English is my job. （不定式作主语） 注意 不定式作主语时，常用形式主语it句型，因此左例可变为It is my job to teach them English. （真正的主语是to teach them English.） 2 谓语 说明主语“做什么”，“是什么”或“怎么样”。 谓语（谓语部分里主要的词）必须用动词。谓语和主语在人称和数两方面必须一致。它在主语后面。 His Parents are doctors. （系动词作谓语） She looks well.（系动词作谓语） We study hard.（实义动词作谓语） He can speak English. （情态动词和实义动词作谓语） 3 表语 表语说明主语“是什么”或者“怎么样”，由名词、形容词、副词、介词、不定式及相当于名词或形容词的词或短语来担任。它的位置在系动词后面。 You look younger than before.（形容词作表语） I am a teacher.（名词作表语） Everybody is here.（副词作表语） They are at home now.（介词短语作表语） My job is to teach them English.（不定式作表语） 4 ,宾语 宾语是动作、行为的对象，由名词、代词、不定式或相当于名词的词、短语来担任，它和及物动词一起说明主语做什么，在谓语之后。

Itsinceitbefore句型和练习

2. It will be two years the economic situation improves. It+be+ 时间 +before/since/when/that 句型辨析 I. since 1.It is/has bee n+ 时间段+si nee sb did sth 某个动作发生持续多久 eg:It is/has been three years since he finished the work. 自从他完成这项工作已经三年了 . 2. 该句型中若 since 后跟延续性动词，要翻译成否定含义 , 即“没做某事已经多 久了”. eg:It has been three years since he worked here. 他不在这工作已经三年 了. It has been three years since he smoked. 他已经戒烟三年了 . 但是：It has been three years since he began to smoke. 他吸烟有三年了。 II .before. 1.It was/will be+ 时间段+before... 过多长时间将发生某动作或事情；或过 了多久发生了某事或动作。 eg:It was three years before he finished the work. 项工作. It will be three years before he finishes the work. 这项工 作 . 2.It won't be/take long before... 不久就会… eg:It won't be long before he finishes the work. 作. III when. when 没有固定与It is …连用的句型.when 可表示时间点或时间段，因此从句 中短暂性动词或延续性动词均可用 .when 还可作并列连词，表示突然发生一个 动作, 等于 and at that time. 常用句型 : 1 主语 +was/were doing whe n... 2 主语 +was/were about to do whe n... It+be+ 时间 +before/si nce/whe n/that 重要考点练习 It +be+时间+before/s in ce/whe n/that 是高考中每年必考的重要考点，而学生却对这个句型 中几个关联词的选择把握不准，造成失分。我在试题讲解的过程中设计了以下几个小题， 让学生去分析，整理，归纳，效果很好。 1. It was two years _______ h e realized the truth. 3. It was two years later _______ he realized the truth. 他花了三年才完成这 他得花三年才能完成 他不久就会完成这项工