Comparison of photobioreactors and optimal light regime for rapid vegetative propagation of transgenic Undaria pinnatifida gametophytes

BACKGROUND: Previously, tachyplesin gene (tac) has been successfully transferred into Undaria pinnatifida gametophytes using the method of microprojectile bombardment transformation. The objectives of this study were to compare and evaluate the performance of bubble-column and airlift bioreactors to determine a preferred configuration of bioreactor for vegetative propagation of transgenic U. pinnatifida gametophytes, and to then investigate the influence of light on vegetative propagation of these gametophytes, including incident light intensity, photoperiod and light quality to resolve the problems of rapid vegetative propagation within the selected bioreactor. RESULTS: Experimental results showed that final dry cell density in the airlift bioreactor was 12.7% higher than that in the bubble-column bioreactor under the optimal aeration rate of 1.2 L air min(-1) L-1 culture. And a maximum final dry cell density of 2830 mg L-1 was obtained within the airlift bioreactor using blue light at 40 mu mol m(-2) s(-1) with a light/dark cycle of 14/10 (h). Polymerase chain reaction (PCR) analysis indicated that genes (bar and tac) were not lost during rapid vegetative propagation within the airlift bioreactor. CONCLUSION: The airlift bioreactor was shown to be much more suitable for rapid vegetative propagation of transgenic U. pinnatifida gametophytes than the bubble-column bioreactor in the laboratory. The use of blue light allows improvement of vegetative propagation of transgenic U. pinnatifida gametophytes. (C) 2009 Society of Chemical Industry

Ontogenetic development of digestive enzymes and effect of starvation in miiuy croaker Miichthys miiuy larvae

The ontogenetic development of the digestive enzymes amylase, lipase, trypsin, and alkaline phosphatase and the effect of starvation in miiuy croaker Miichthys miiuy larvae were studied. The activities of these enzymes were detected prior to exogenous feeding, but their developmental patterns differed remarkably. Trypsin activity continuously increased from 2 days after hatching (dah), peaked on 20 dah, and decreased to 25 dah at weaning. Alkaline phosphatase activity oscillated at low levels within a small range after the first feeding on 3 dah. In contrast, amylase and lipase activities followed the general developmental pattern that has been characterized in fish larvae, with a succession of increases or decreases. Amylase, lipase, and trypsin activities generally started to increase or decrease at transitions from endogenous to exogenous feeding or diet changes, suggesting that these enzymatic activities can be modulated by feeding modes. The activities of all the enzymes remained stable from 25 dah onwards, coinciding with the formation of gastric glands and pyloric caecum. These results imply that specific activities of these enzymes underwent changes due to morphological and physiological modifications or diet shift during larval development but that they became stable after the development of the digestive organs and associated glands was fully completed and the organs/glands functioned. Trypsin and alkaline phosphatase were more sensitive to starvation than amylase and lipase because delayed feeding up to 2 days after mouth opening was able to adversely affect their activities. Enzyme activities did not significantly differ among feeding groups during endogenous feeding; however, all activities were remarkably reduced when delayed feeding was within 3 days after mouth opening. Initiation of larvae feeding should occur within 2 days after mouth opening so that good growth and survival can be obtained in the culture.

Suspension culture of gametophytes of transgenic kelp in a photobioreactor

Transgenic Laminaria japonica gametophytes producing a recombinant tissue-type plasminogen activator (rtPA) protein, which is an effective third-generation thrombolytic agent for acute myocardial infarction (AMI), were cultured in an illuminated bubble column bioreactor. A maximum final dry cell weight of 1120 mg l(-1) was obtained in batch culture with an initial dry cell weight of 126 mg l(-1) and with aeration rate of 1.2 l air min(-1) l(-1) culture, nitrate at 1.5 mM and phosphate at 0.17 mM. The yield of rtPA was 56 mu g g(-1) dry cell wt. This is the first report regarding cultivation of a transgenic macroalga in a bioreactor.

Three-stage transformation of chlorophyll transient fluorescence pattern under sustained dehydration and the discovery of critical water content in seaweeds

The chlorophyll fluorescence kinetics of marine red alga Grateloupia turutunt Yamada, green alga Ulva pertusa Kjellm and brown alga Laminaria japonica Aresch during natural sustained dehydration were monitored and investigated. The pulse amplified modulation (PAM) system was used to analyze the distinct fluorescence parameters during thallus dehydration. Results proved that the fluorescence kinetics of different seaweed all showed three patterns of transformation with sustained water loss. These were: 1) peak kinetic pattern (at the early stage of dehydration fluorescence enhanced and quenched subsequently, representing a normal physiological state). 2) plateau kinetic pattern (with sustained water loss fluorescence enhanced continuously but quenching became slower, finally reaching its maximum). 3) Platform kinetic pattern (fluorescence fell and the shape of kinetic curve was similar to plateau kinetic pattern). A critical water content (CWC) could be found and defined as the percentage of water content just prior to the fluorescence drop and to be a significant physiological index for evaluation of plant drought tolerance. Once thallus water content became lower than this value the normal peak pattern can not be recovered even through rehydration, indicating an irreversible damage to the thylakoid membrane. The CWC value corresponding to different marine species were varied and negatively correlated with their desiccation tolerance, for example. Laminaria japonica had the highest CWC value (around 90%) and the lowest dehydration tolerance of the three. In addition, a fluorescence "burst" was found only in red algae during rehydration. The different fluorescence parameters F-o, F-v and F-v, F-m were measured and compared during water loss. Both F-o and F-v increased in the first stage of dehydration but F-v/F-m. kept almost constant. So the immediate response of in vivo chlorophyll fluorescence to dehydration was an enhancement. Later with sustained dehydration F-o increased continuously while F-v decreased and tended to become smaller and smaller. The major changes in fluorescence (including fluorescence drop during dehydration and the burst during rehydration) were all attributed to the change in F-o instead of F-v This significance of F-o indicates that it is necessary to do more research on F-o as well as on its relationship with the state of thylakoid membrane.

Early expressed genes showing a dichotomous developing pattern in the lancelet embryo

Lancelets (amphioxus), although showing the most similar anatomical features to vertebrates, never develop a vertebrate-like head but rather several structures specific to this animal. The lancelet anatomical specificity seems to be traceable to early developmental stages, such as the vertebrate dorsal and anterior-posterior determinations. The BMP and Wnt proteins play important roles in establishing the early basis of the dorsal structures and the head in vertebrates. The early behavior of BMP and Wnt may be also related to the specific body structures of lancelets. The expression patterns of a dpp-related gene, Bbbmp2/4, and two wnt-related genes, Bbwnt7 and Bbwnt8, have been studied in comparison with those of brachyury and Hnf-3 beta class genes The temporal expression patterns of these genes are similar to those of vertebrates; Bbbmp2/4 and Bbwnt8 are first expressed in the invaginating primitive gut and the equatorial region. respectively, at the initial gastrula stage. However, spatial expression pattern of Bbbmp2/4 differs significantly from the vertebrate cognates. It is expressed in the mid-dorsal inner layer of gastrulae and widely in the anterior region, in which vertebrates block BMP signaling, The present study suggests that the lancelet embryo may have two distinct developmental domains from the gastrula stage, the domains of which coincide later with the lateral diverticular and the somitocoelomic regions. The embryonic origin of the anterior-specific structures in lancelets corresponds to the anterior domain where Bbbmp2/4 is continuously expressed.

Expression, purification, and characterization of recombinant Chinese shrimp crustin-like protein (CruFc) in Pichia pastoris

A crustin-like protein (CruFc) from Fenneropenaeus chinensis was expressed in Pichia pastoris and then purified to electrophoretic homogeneity on a Sephacryl S-100 column with a band corresponding to the expected one (13 kDa) shown by 15% SDS-PAGE. Western blot indicated that the rCruFc specifically reacted with polyclonal rabbit anti-Fenneropenaeus chinensis CruFc. Production in a 5 l bioreactor gave 237 mg rCruFc/l. Antimicrobial assay revealed that 4 mu M rCruFc inhibited growth of Staphylococcus aureus.

Recombinant expression of rt-PA gene (encoding Reteplase) in gametophytes of the seaweed Laminaria japonica (Laminariales, Phaeophyta)

The life cycle of seaweed Laminaria japonica involves a generation alternation between diploid sporophyte and haploid gametophte. The expression of foreign genes in sporophte has been proved. In this research, the recombinant expression in gametophyte was investigated by particle bombardment with the rt-PA gene encoding the recombinant human tissue-type plasminogen activator (Reteplase), which is a thrombolytic agent for acute myocardial infarction (AMI). Transgenic gametophytes were selected by their resistance to herbicide phosphiothricin (PPT), and proliferated in an established bubble column photo-bioreactor. According to the results from quantitative ELISA, Southern blotting, and fibrin agarose plate assay (FAPA) for bioactivity, it was showed that the rt-PA gene had been integrated into the genome of gametophytes of L. japonica, and the expression product showed the expected bioactivity, implying the proper post-transcript modification in haploid gametophyte.

Distribution and their pollution assessment of heavy metals in the sediments of the Yalu River Estuary and its adjacent coastal waters

The present paper deals with the distribution patterns of heavy metals and the associated influencing factors in the Yalu River Estuary and its adjacent coastal waters. Based upon the analysis of the surficial and core sediments measurements, the pollution of heavy metal and potential ecological risk were evaluated. The burial flux and contents of heavy metals (except for copper) have been continuously increasing since the 1920s. Therefore, the gross potential ecological risk for the sediments was high or very high, and the study area was endangered by heavy metals contamination. Heavy metals originated mainly from upstream pollutant input, correlation analysis showed that chromium, nickel, zinc, cadmium, lead, arsenic, and mercury in the sediments of the middle and west channels as well as the sea area of the western Yalu River Estuary concentrations were most probably derived from similar sources. In contrast, the metal of copper most probably originated from sources different from the other metals. Preliminary studies indicate that copper contamination was most likely the result of emission from mining activities situated at the upstream of the river. The contents of heavy metals in the sediments of estuarine turbidity maximum zone of Yalu River were larger than those of any other areas in the middle channel. With large portion of fine sediments, weaker hydrodynamics, and richer sources of heavy metals, the sediments of the west channel, were even more enriched with heavy metals than those of the middle channel.

Dynamics of intraseasonal sea level and thermocline variability in the equatorial Atlantic during 2002-03

Satellite and in situ observations in the equatorial Atlantic Ocean during 2002-03 show dominant spectral peaks at 40-60 days and secondary peaks at 10-40 days in sea level and thermocline within the intraseasonal period band (10-80 days). A detailed investigation of the dynamics of the intraseasonal variations is carried out using an ocean general circulation model, namely, the Hybrid Coordinate Ocean Model (HYCOM). Two parallel experiments are performed in the tropical Atlantic Ocean basin for the period 2000-03: one is forced by daily scatterometer winds from the Quick Scatterometer (QuikSCAT) satellite together with other forcing fields, and the other is forced by the low-passed 80-day version of the above fields. To help in understanding the role played by the wind-driven equatorial waves, a linear continuously stratified ocean model is also used. Within 3 degrees S-3 degrees N of the equatorial region, the strong 40-60-day sea surface height anomaly (SSHA) and thermocline variability result mainly from the first and second baroclinic modes equatorial Kelvin waves that are forced by intraseasonal zonal winds, with the second baroclinic mode playing a more important role. Sharp 40-50-day peaks of zonal and meridional winds appear in both the QuikSCAT and Pilot Research Moored Array in the Tropical Atlantic (PIRATA) data for the period 2002-03, and they are especially strong in 2002. Zonal wind anomaly in the central-western equatorial basin for the period 2000-06 is significantly correlated with SSHA across the equatorial basin, with simultaneous/ lag correlation ranging from-0.62 to 0.74 above 95% significance. Away from the equator (3 degrees-5 degrees N), however, sea level and thermocline variations in the 40-60-day band are caused largely by tropical instability waves (TIWs). On 10-40-day time scales and west of 10 degrees W, the spectral power of sea level and thermocline appears to be dominated by TIWs within 5 degrees S-5 degrees N of the equatorial region. The wind-driven circulation, however, also provides a significant contribution. Interestingly, east of 10 W, SSHA and thermocline variations at 10 40- day periods result almost entirely from wind-driven equatorial waves. During the boreal spring of 2002 when TIWs are weak, Kelvin waves dominate the SSHA across the equatorial basin (2 degrees S-2 degrees N). The observed quasi-biweekly Yanai waves are excited mainly by the quasi-biweekly meridional winds, and they contribute significantly to the SSHA and thermocline variations in 1 degrees-5 degrees N and 1 degrees-5 degrees S regions.

Analysis of the sea-level variability along the Chinese coast and estimation of the impact of a CO2-perturbed atmospheric circulation

The relationship between monthly sea-level data measured at stations located along the Chinese coast and concurrent large-scale atmospheric forcing in the period 1960-1990 is examined. It is found that sea-level varies quite coherently along the whole coast, despite the geographical extension of the station set. A canonical correlation analysis between sea-level and sea-level pressure (SLP) indicates that a great part of the sea-level variability can be explained by the action of the wind stress on the ocean surface. The relationship between sea-level and sea-level pressure is analyzed separately for the summer and winter half-years. In winter, one factor affecting sea-level variability at all stations is the SLP contrast between the continent and the Pacific Ocean, hence the intensity of the winter Monsoon circulation. Another factor that affects coherently all stations is the intensity of the zonal circulation at mid-latitudes. In the summer half year, on the other hand, the influence of SLP on sea-level is spatially less coherent: the stations in the Yellow Sea are affected by a more localized circulation anomaly pattern, whereas the rest of the stations is more directly connected to the intensity of the zonal circulation. Based on this analysis, statistical models (different for summer and winter) to hindcast coastal sealevel anomalies from the large-scale SLP field are formulated. These models have been tested by fitting their internal parameters in a test period and reproducing reasonably the sea-level evolution in an independent period. These statistical models are also used to estimate the contribution of the changes of the atmospheric circulation on sea-level along the Chinese coast in an altered climate. For this purpose the ouput of 150 year-long experiment with the coupled ocean-atmosphere model ECHAM1-LSG has been analyzed, in which the atmospheric concentration of greenhouse gases was continuously increased from 1940 until 2090, according to the Scenario A projection of the Intergovermental Panel on Climate Change. In this experiment the meridional (zonal) circulation relevant for sea-level tends to become weaker (stronger) in the winter half year and stronger (weaker) in summer. The estimated contribution of this atmospheric circulation changes to coastal sea-level is of the order of a few centimeters at the end of the integration, being in winter negative in the Yellow Sea and positive in the China Sea with opposite signs in the summer half-year.

STUDY ON THE FORMATIONAL MECHANISM OF THE NORTHERN YELLOW (HUANGHAI) SEA COLD WATER MASS .2. DISCUSSION ON THE SOLUTION OF THE MODEL

The theoretical solution of the model of the Northern Yellow (Huanghai) Sea Cold Water Mass (NYSCWM) reveals that the NYSCWM is mainly formed through the continuous temperature increase of the overwintered water body above the Northern Yellow Sea Depression (NYSD) after spring when heat is continuously conducted from the sea surface to the deeper layer. In the NYSCWM's growing period, (June-July), nonlinear vertical convection and advection effects continuously increase, and are gradually balanced by the heat diffusion effect as the temperature increases from the surface to the bottom, which leads to the formation of an intensive thermocline and lateral front. Meanwhile, the three-dimensional circulation correspondingly occurs. In the NYSCWM's entire growing period, the horizontal circulation is always in the cyclonic motion, while the vertical circulation passes through a transition from a period with the cold centre as downwelling to a period with the cold centre as upwelling.

淤泥质潮滩地貌的遥感研究－以苏北辐射沙洲海岸为例

本文以具有典型特征的苏北淤泥质潮滩海岸作为研究区，利用1975-2003年间14景覆盖该地区的Landsat和SPOT卫星影像作为主要数据源，结合地面调查和验证工作，在遥感影像处理和地理信息系统分析技术的支持下，对区内潮滩、岸线、水边线和盐沼植被等进行遥感解译，分析苏北辐射沙脊群和沿岸地貌的空间分布特征和动态演变趋势。研究结果表明：苏北辐射沙脊群海域的潮汐水位过程的不同步现象普遍存在，限制了常规遥感数据在苏北潮滩地貌研究中的适用范围和解译精度；在人工判别的辅助下，多光谱遥感的非监督分类方法可以有效解译淤泥质潮滩的水边线；利用修改型土壤调整植被指数（MSAVI）可以较好地提取潮滩上的盐沼植被信息；苏北沿岸潮滩的快速淤长促进了盐沼植被带向海侧快速扩展，近年来持续的潮滩围垦工程则不断从陆侧侵占盐沼植被带，使盐沼植被带宽度减小乃至消失；在大规模人类活动和自然条件的共同影响下，苏北辐射沙脊群海岸的岸线发育趋于平直化，无序的潮滩围垦项目使得可垦滩地资源被过度消耗；1975～2002年间，研究区北部和南部沿岸的高潮滩整体上处于淤长状态，中部沿岸潮滩和离岸沙洲高潮滩则被大面积侵蚀；1999年以来，研究区内低潮滩部位开始形成有序排列的滩面地物，并表现出逐年大面积蔓延的趋势，可能是滩涂紫菜养殖区扩展的结果。

南海北部陆缘深水区构造演化及其资源效应

南海北部陆缘深水区（水深>300m）蕴藏着丰富的资源，我国对深水区的地质研究刚刚起步，但相关领域已成为科研热点。深水油气盆地的构造演化是油气勘探中最重要的基础性研究之一，因此针对我国南海北部陆缘深水区开展构造演化及其资源效应的研究具有重要的理论意义和实际意义。 本文利用钻井和地震资料并结合区域地质资料，重点研究了珠江口盆地深水区的结构和构造演化，取得如下创新性成果：1）首次利用半地堑分析方法系统解剖了研究区的结构、各构造单元发育特征，在此基础上指出五个有利油气运聚带；2）采用回剥法并利用最新资料进行校正，得到了研究区更为可靠的构造沉降曲线，重新划分了裂陷期和裂后期的分界，认为32Ma南海海底扩张之后裂陷作用仍在持续，直到23Ma左右才开始大规模裂后热沉降，并进一步解释了裂陷期延迟的形成机制；3）应用非连续拉张模型计算拉张系数的方程计算了研究区的壳幔拉张系数，指出了深水区地幔相对于地壳的优势伸展作用；首次运用平衡剖面技术重建了研究区的构造发育史，计算了各构造期的拉张率和沉积速率，指出研究区新生代整体呈现持续拉张，拉张系数在1.1-1.24之间；4）精细刻画了水合物钻采区的地质构造特征，建立了该区天然气水合物成藏的概念模式；建立了一套根据地震叠加速度计算流体势的方法，为水合物成藏规律的研究提供了新的思路。

Effect of water temperature on digestive enzyme activity and gut mass in sea cucumber Apostichopus japonicus (Selenka), with special reference to aestivation

The effect of water temperature on gut mass and digestive enzyme activity in sea cucumber Apostichopus japonicus, including relative gut mass (RGM), amylase, lipase, pepsin and trypsin activities were studied at temperatures of 7, 14, 21, and 28A degrees C over a period of 40 days. Results show that RGM significantly decreased after 40 days at 21A degrees C and markedly decreased over the whole experiment period at 28A degrees C; however, no significant effect of duration was observed at 7 or 14A degrees C. At 14A degrees C, trypsin activity significantly decreased over 10 and 20 days, then increased; amylase and trypsin activity significantly decreased after 40 days at 28A degrees C. However, no significant effect of duration was found on amylase, pepsin or trypsin activities in the other temperature treatment groups. At 28A degrees C, lipase activity peaked in 20 days and then markedly decreased to a minimum at the end of the experiment. On the other hand, pepsin activity at 28A degrees C continuously increased over the whole experimental period. Principle component analysis showed that sea cucumbers on day 40 in the 21A degrees C group and in the previous 20 days in the 28A degrees C group were in the prophase of aestivation. At 28A degrees C, sea cucumbers aestivated at 30-40 days after the start of the experiment. It is concluded that the effect of temperature on the digestion of A. japonicus is comparatively weak within a specific range of water temperatures and aestivation behavior is accompanied by significant changes in RGM and digestive enzyme activities.

Forms and functions of inorganic carbon in the Jiaozhou Bay sediments

Inorganic carbon forms and their influencing factors, mutual transformation and contribution to carbon cycling in the Jiaozhou Bay sediments were discussed. The results show that inorganic carbon in sediments could be divided into five forms: NaCl form, NH3 center dot H2O form, NaOH form, NH2OH center dot HCl form and HCl form. Thereinto, NH2OH center dot HCl form and HCl form account for more than 70% of total inorganic carbon. There was close relationship among every form of inorganic carbon and their correlativity was clearly different with different sedimentary environment except the similar strong positive correlation among NH2OH center dot HCl form, HCl form and total inorganic carbon in all regions of the Jiaozhou Bay. All forms of inorganic carbon were influenced by organic carbon, pH, Eh, Es, nitrogen and phosphorus in sediments, but their influence had different characteristics in different regions. Every farm of inorganic carbon transformed into each other continuously during early diagenesis of sediments and the common phenomenon was that NaCl form, NH3 center dot H2O form, NaOH form and NH2OH center dot HCl form might transform into steady HCl form. NaCl form, NH3 center dot H2O form, NaOH form and NH2OH center dot HCl form could participate in carbon recycle and they are potential carbon source; HCl form may be buried for a long time in sediments, and it may be one of the final resting places of atmospheric CO2. Inorganic carbon which entered into sediments was about 4.98 x 10(10) g in the Jiaozhou Bay every year, in which about 1.47x10(10) g of inorganic carbon might be buried for a long time and about 3.51. x 10(10) g of inorganic carbon might return into seawater and take part in carbon recycling.