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61.
Zaharoff DA Henshaw JW Mossop B Yuan F 《Experimental biology and medicine (Maywood, N.J.)》2008,233(1):94-105
Pulsed electric field has been widely used as a nonviral gene delivery platform. The delivery efficiency can be improved through quantitative analysis of pore dynamics and intracellular transport of plasmid DNA. To this end, we investigated mechanisms of cellular uptake of macromolecules during electroporation. In the study, fluorescein isothiocyanate-labeled dextran (FD) with molecular weight of 4,000 (FD-4) or 2,000,000 (FD-2000) was added into suspensions of a murine mammary carcinoma cell (4T1) either before or at different time points (ie, 1, 2, or 10 sec) after the application of different pulsed electric fields (in high-voltage mode: 1.2-2.0 kV in amplitude, 99 microsec in duration, and 1-5 pulses; in low-voltage mode: 100-300 V in amplitude, 5-20 msec in duration, and 1-5 pulses). The intracellular concentrations of FD were quantified using a confocal microscopy technique. To understand transport mechanisms, a mathematical model was developed for numerical simulation of cellular uptake. We observed that the maximum intracellular concentration of FD-2000 was less than 3% of that in the pulsing medium. The intracellular concentrations increased linearly with pulse number and amplitude. In addition, the intracellular concentration of FD-2000 was approximately 40% lower than that of FD-4 under identical pulsing conditions. The numerical simulations predicted that the pores larger than FD-4 lasted <10 msec after the application of pulsed fields if the simulated concentrations were on the same order of magnitude as the experimental data. In addition, the simulation results indicated that diffusion was negligible for cellular uptake of FD molecules. Taken together, the data suggested that large pores induced in the membrane by pulsed electric fields disappeared rapidly after pulse application and convection was likely to be the dominant mode of transport for cellular uptake of uncharged macromolecules. 相似文献
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In China, croplands account for a relatively large form of vegetation cover. Quantifying carbon dioxide exchange and understanding the environmental controls on carbon fluxes over croplands are critical in understanding regional carbon budgets and ecosystem behaviors. In this study, the net ecosystem exchange (NEE) at a winter wheat/summer maize rotation cropping site, representative of the main cropping system in the North China Plain, was continuously measured using the eddy covariance technique from 2005 to 2009. In order to interpret the abiotic factors regulating NEE, NEE was partitioned into gross primary production (GPP) and ecosystem respiration (Reco). Daytime Reco was extrapolated from the relationship between nighttime NEE and soil temperature under high turbulent conditions. GPP was then estimated by subtracting daytime NEE from the daytime estimates of Reco. Results show that the seasonal patterns of the temperature responses of Reco and light‐response parameters are closely related to the crop phenology. Daily Reco was highly dependent on both daily GPP and air temperature. Interannual variability showed that GPP and Reco were mainly controlled by temperature. Water availability also exerted a limit on Reco. The annual NEE was ?585 and ?533 g C m?2 for two seasons of 2006–2007 and 2007–2008, respectively, and the wheat field absorbed more carbon than the maize field. Thus, we concluded that this cropland was a strong carbon sink. However, when the grain harvest was taken into account, the wheat field was diminished into a weak carbon sink, whereas the maize field was converted into a weak carbon source. The observations showed that severe drought occurring during winter did not reduce wheat yield (or integrated NEE) when sufficient irrigation was carried out during spring. 相似文献
64.
RACE技术是一项扩增基因末端序列的新技术。该研究从牛BMP4基因出发,以牛软骨的RNA为模板,按照不同物种BMP4基因的相似性设计特异引物,运用PCR和RACE技术扩增并获得了特异片段,该片段经PCR、酶切和测序验证,证实所克隆序列为牛BMP4的3′端序列,包含有1170bp组成的开放读码框(ORF),编码389个氨基酸,3′非编码区121bp个核苷酸和poly(A)15。同源性分析结果表明,牛BMP4 cDNA最大开放读码框所推测的氨基酸序列与已知人、小鼠、大鼠、狗、羊和鸡等真核生物BMP4氨基酸序列进行比较,分别有94.5%、93.1%、91.9%、87.4%、94.2%、79%的同源性。这为克隆其他物种的BMP4基因提供了依据,同时牛骨形态发生蛋白的测序为我们更好的理解牛的生骨机理提供帮助。 相似文献
65.
目的: 真核细胞表达小鼠淋巴细胞抗原CTLA-4胞外段肽,研究表达肽段与抗原呈递细胞B7分子结合后减轻小鼠淋巴细胞刺激后的增殖抑制,从而启动T淋巴细胞进一步增殖。方法:从小鼠脾脏淋巴细胞获得总RNA,通过逆转录PCR扩增出CTLA-4全长基因,克隆并测序。依据胞外段序列和真核表达载体pcDNA3.1序列,合成引物扩增胞外片段,两者经内切核酸酶处理、连接构建重组表达pcDNA3.1载体,重组质粒经测序验证后,采用lipofectamine 2000转染入小鼠肝癌细胞Hepa1-6,经G418筛选获得稳定表达细胞株。结果:获得小鼠CTLA-4胞外段真核表达载体和小鼠肝癌细胞Hepa1-6稳定表达转染细胞株,制备了CTLA-4胞外肽段,经His标签抗体和小鼠CTLA-4抗体Western blot检测表达蛋白带均呈阳性。结论:获得CTLA-4胞外段肽,为进一步研究该肽的作用打下基础。 相似文献
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MÁRCIA FERNANDES DE AQUINO SANTOS JOSÉ RICARDO M. MERMUDES VERA MARIA MEDINA DA FONSECA 《Palaeontology》2011,54(4):807-814
Abstract: A specimen of Curculionidae (Curculioninae) is described as Arariperhinus monnei gen. et sp. nov. The specimen is preserved on a laminated limestone sample of the Crato Formation (Santana Group), Lower Cretaceous (Aptian–Albian), and was collected from a quarry near Nova Olinda, Chapada do Araripe, State of Ceará, Brazil. The genus is placed in the subfamily Curculioninae because of its strongly convex body and relatively slender rostrum, but mainly by its rounded eyes and lack of a prosternal sulcus and tibial spurs. The very prominent eyes in lateral view, a cylindrical rostrum and a straight posterior margin of ventrite II are strong indications that this fossil belongs to the tribe Anthonomini. However, the claws, which would resolve the exact placement of this fossil, are poorly preserved. Arariperhinus monnei gen. et sp. nov. is distinguishable by the combination of several characters and the first record of the family Curculionidae in the Santana Group; it is the oldest record of a member of the subfamily Curculioninae. 相似文献
68.
RICARDO CRUZ DE CARVALHO ANABELA BERNARDES DA SILVA RENATA SOARES ANDRÉ M. ALMEIDA ANA VARELA COELHO JORGE MARQUES DA SILVA CRISTINA BRANQUINHO 《Plant, cell & environment》2014,37(7):1499-1515
All bryophytes evolved desiccation tolerance (DT) mechanisms during the invasion of terrestrial habitats by early land plants. Are these DT mechanisms still present in bryophytes that colonize aquatic habitats? The aquatic bryophyte Fontinalis antipyretica Hedw. was subjected to two drying regimes and alterations in protein profiles and sucrose accumulation during dehydration and rehydration were investigated. Results show that during fast dehydration, there is very little variation in protein profiles, and upon rehydration proteins are leaked. On the other hand, slow dehydration induces changes in both dehydration and rehydration protein profiles, being similar to the protein profiles displayed by the terrestrial bryophytes Physcomitrella patens (Hedw.) Bruch and Schimp. and, to what is comparable with Syntrichia ruralis (Hedw.) F. Weber and D. Mohr. During dehydration there was a reduction in proteins associated with photosynthesis and the cytoskeleton, and an associated accumulation of proteins involved in sugar metabolism and plant defence mechanisms. Upon rehydration, protein accumulation patterns return to control values for both photosynthesis and cytoskeleton whereas proteins associated with sugar metabolism and defence proteins remain high. The current results suggest that bryophytes from different ecological adaptations may share common DT mechanisms. 相似文献
69.
Graham RD Jones Sabrina DA Koetsier 《The Clinical biochemist. Reviews / Australian Association of Clinical Biochemists》2014,35(4):243-250
Reference intervals are commonly considered to allow for between-laboratory bias. The RCPAQAP Liquid Serum Chemistry Program has collected data on laboratory measurements as well as reference intervals. This allows assessment of the between-laboratory variation in results, reference intervals and the information transmitted by the combination of these factors. For the majority of common chemistry analytes, the between-laboratory variation in reference intervals is greater than the variation in results. Additionally the reference interval variation is generally not related to bias between the results. Use of common reference intervals, either as an average of the current intervals in use, or the intervals proposed by the AACB Harmonisation Group, improved the variation seen in the information produced by different laboratories. 相似文献
70.