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Recombinant adeno-associated virus(rAAV) has proven to be a promising gene delivery vector for human gene therapy. However, its application has been limited by difficulty in obtaining enough quantities of high-titer vector stocks. In this paper, a novel and highly efficient production system for rAAV is described. A recombinant herpes simplex virus type 1(rHSV-1) designated HSV1-rc/△UL2, which expressed adeno-associated virus type2(AAV-2) Rep and Cap proteins, was constructed previously. The data confirmed that its functions were to support rAAV replication and packaging, and the generated rAAV was infectious. Meanwhile, an rAAV proviral cell line designated BHK/SG2, which carried the green fluorescent protein(GFP) gene expression cassette, was established by transfecting BHK-21 cells with rAAV vector plasmid pSNAV-2-GFP. Infecting BHK/SG2 with HSV1-rc/△UL2 at an MOI of 0.1 resulted in the optimal yields of rAAV, reaching 250 transducing unit(TU) or 4.28×104 particles per cell. Therefore, compared with the conventional transfection method, the yield of rAAV using this "one proviral cell line, one helper virus" strategy was increased by two orders of magnitude. Large-scale production of rAAV can be easily achieved using this strategy and might meet the demands for clinical trials of rAAV-mediated gene therapy. 相似文献
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The Tibetan Plateau is an essential area to study the potential feedback effects of soils to climate change due to the rapid rise in its air temperature in the past several decades and the large amounts of soil organic carbon (SOC) stocks, particularly in the permafrost. Yet it is one of the most under-investigated regions in soil respiration (Rs) studies. Here, Rs rates were measured at 42 sites in alpine grasslands (including alpine steppes and meadows) along a transect across the Tibetan Plateau during the peak growing season of 2006 and 2007 in order to test whether: (1) belowground biomass (BGB) is most closely related to spatial variation in Rs due to high root biomass density, and (2) soil temperature significantly influences spatial pattern of Rs owing to metabolic limitation from the low temperature in cold, high-altitude ecosystems. The average daily mean Rs of the alpine grasslands at peak growing season was 3.92 μmol CO(2) m(-2) s(-1), ranging from 0.39 to 12.88 μmol CO(2) m(-2) s(-1), with average daily mean Rs of 2.01 and 5.49 μmol CO(2) m(-2) s(-1) for steppes and meadows, respectively. By regression tree analysis, BGB, aboveground biomass (AGB), SOC, soil moisture (SM), and vegetation type were selected out of 15 variables examined, as the factors influencing large-scale variation in Rs. With a structural equation modelling approach, we found only BGB and SM had direct effects on Rs, while other factors indirectly affecting Rs through BGB or SM. Most (80%) of the variation in Rs could be attributed to the difference in BGB among sites. BGB and SM together accounted for the majority (82%) of spatial patterns of Rs. Our results only support the first hypothesis, suggesting that models incorporating BGB and SM can improve Rs estimation at regional scale. 相似文献
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Vascular structures in natural systems are able to provide high mass transport through high surface areas and optimized structure. Few synthetic material fabrication techniques are able to mimic the complexity of these structures while maintaining scalability. The Vaporization of a Sacrificial Component (VaSC) process is able to do so. This process uses sacrificial fibers as a template to form hollow, cylindrical microchannels embedded within a matrix. Tin (II) oxalate (SnOx) is embedded within poly(lactic) acid (PLA) fibers which facilitates the use of this process. The SnOx catalyzes the depolymerization of the PLA fibers at lower temperatures. The lactic acid monomers are gaseous at these temperatures and can be removed from the embedded matrix at temperatures that do not damage the matrix. Here we show a method for aligning these fibers using micromachined plates and a tensioning device to create complex patterns of three-dimensionally arrayed microchannels. The process allows the exploration of virtually any arrangement of fiber topologies and structures. 相似文献
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2011年4和8月及2014年4月在陕西省宝鸡市太白县分别采集到2只、1只和1只共4只蝙蝠个体,标本保存于陕西省动物研究所标本馆(标本编号为TB0001、TB0002、TB0017和TB0042)。其主要形态特征为:体型小,头体长34.59 ~ 43.86 mm(n = 4);耳较小,黑色;耳前缘近凸圆,耳后缘在基部有一浅的缺刻;耳屏细长,约为耳长的一半;后足不及胫长的一半;背部毛基黑色,毛尖为赤褐色;腹部毛基为黑色,毛尖为灰褐色;翼膜起始于后足外趾的基部。以上特征均与喜山鼠耳蝠moupinensis亚种(Myotis muricola moupinensis)相符。结合线粒体Cyt b基因序列,构建其系统发生关系,与基因库中宽吻蝠属未定种(Submyotodon sp.)序列聚为较高支持度的一支。综合形态与系统发育证据,参考目前最新的中国兽类物种分类和分布信息,将采集到的标本初步鉴定为喜山鼠耳蝠moupinensis亚种,此次发现是陕西省境内该物种的首次记录,拓展了对其在我国分布范围的认识。 相似文献
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转基因红花中角质细胞生长因子KGF-1的表达 总被引:3,自引:0,他引:3
通过构建重组表达质粒载体p139035S-KGF1和根癌农杆菌介导在红花(Carthamus tinctorius)中表达角质细胞生长因子(KGF-1)。从侵染到诱导生根共需要14周, 转化率达0.1%。红花子叶在潮霉素筛选培养基上培养4–5周后便可获得丛生芽, 再生芽移入含潮霉素的伸长生根培养基, 培养4–8周可诱导生根。通过PCR、Southern blot、RT-PCR及Western blot检测证明目的基因KGF-1已经整合到红花细胞的染色体中, 实现了KGF-1外源蛋白在红花中的成功表达, 为开发KGF-1蛋白新的生产途径奠定了基础。 相似文献
40.
A monoclonal antibody produced by hydridoma cell line, ATCC HB8209, was used to detect and purify erythropoietin synthesized in a cell-free system. The antibody was raised against the N-terminal 20 residues of erythropoietin. It retained anti-erythropoietin activity in 6 M urea in which most of the cell-free synthesized erythropoietin became soluble and gave an enhanced activity of the antibody. 相似文献