马铃薯Y病毒外壳蛋白基因的克隆及序列分析

本文报道应用聚合酶链式反应(PCR)技术,在体外扩增马铃薯 Y 病毒外壳蛋白基因及其克隆和序列分析的结果。病毒 RNA 从马铃薯 Y 病毒感染的烟草叶片中提取,用合成的PCR 3引物及 AMV 逆转录酶合成了单链的 cDNA。利用 PCR 技术,经30个循玎的扩增。得到了一特异的0.8kb 片段。克隆后对此片段进行了限制性内切酶物理图谱分析,并测定了其全序列。实验结果证明,我们克隆到的是完整的马铃薯 Y 病毒的外壳蛋白基因。与国外报道的马铃薯 Y 病毒 N 株相比,其核苷酸序列及推测的氨基酸序列的同源率分别为97.8%和97%。将该基因导入马铃薯以期获得抗 Y 病毒马铃薯的工作正在进行。本文还对 PCR 技术用于扩增植物 RNA 病毒的方法以及用基因工程方法培育抗病毒作物新品种的可行性等进行了讨论。     

The use of streptavidin-biotinylglycans as a tool for characterization of oligosaccharide-binding specificity of lectin.

A new rapid and sensitive method for characterizing lectin specificity using streptavidin-biotinylglycans as a tool is presented. This assay is analogous to enzyme immunoassay and takes advantage of the strong, irreversible adsorption of streptavidin to the wells of the chambers of titer plates. A series of streptavidin-biotinylglycans was first coated on a microtiter plate, and then one of six lectins, concanavalin A, wheat germ agglutinin, Phaseolus vulgaris (red kidney bean) erythro-agglutinin, Lens culinaris (lentil) agglutinin, Datura stramoniun agglutinin, or Sambucus nigra (elderberry bark) agglutinin coupled to horseradish peroxidase, was added. After incubation and thorough washing, only the lectin bound to a complementary glycan remained and could be detected and quantified by the peroxidase reaction. It was established that the lectins retained their oligosaccharide-binding specificities after coupling to the peroxidase, that the binding was inhibited by addition of the corresponding sugar inhibitors, and that the color intensity produced by the enzyme reaction is proportional to the amount of lectin-peroxidase bound to biotinylglycan complexed with streptavidin immobilized on the plate. As an example, it was found that the peroxidase-D. stramoniun agglutinin conjugate strongly bound biotinylglycans, GlcNAc3-Man5-R, GalGlcNAc3Man5-R, and GlcNAc3-4Man3-R (R = GlcNAc2-[6-(biotinamido)hexanoyl]-Asn). As little as 10 pmol/ml of lectin was detected. With the growing availability of biotinylglycans, the method should represent a reliable and simple procedure for screening lectin-oligosaccharide recognition qualitatively and quantitatively.     

慢性缺氧对肺动脉内皮依赖性和非内皮依赖性舒张反应的影响

本实验用生物测定法观察了模拟海拔5000m高度连续缺氧20d对大鼠肺动脉内皮依赖性和非内皮依赖性舒张反应的影响。结果显示慢性缺氧明显抑制了肺内和肺外动脉对乙酰胆碱、ATP、硝普钠和异丙肾上腺素舒张反应的敏感性和反应性。在浓度为10~(-6)mol/L时,缺氧组大鼠肺内动脉对乙酰胆碱、硝普钠和异丙肾上腺素的舒张反应分别只有对照组大鼠的61.3％、75.9％和61.7％,对浓度为1.8×10~(-5)mol/L的ATP的舒张反应只有对照组大鼠的64.9％。研究表明,ATP和乙酰胆碱主要是通过内皮舒张因子使肺动脉产生内皮依赖性舒张反应,硝普钠和异丙肾上腺素分别通过直接激活血管平滑肌细胞鸟苷酸环化酶和β受体使血管产生非内皮依赖性舒张反应。缺氧同时抑制了肺动脉内皮依赖性和非内皮依赖性舒张反应,提示慢性缺氧可能抑制了正常肺内舒血管活性物质的生理作用。     

Chaperonins GroEL and GroES: views from atomic force microscopy.

The Escherichia coli chaperonins, GroEL and GroES, as well as their complexes in the presence of a nonhydrolyzable nucleotide AMP-PNP, have been imaged with the atomic force microscope (AFM). We demonstrate that both GroEL and GroES that have been adsorbed to a mica surface can be resolved directly by the AFM in aqueous solution at room temperature. However, with glutaraldehyde fixation of already adsorbed molecules, the resolution of both GroEL and GroES was further improved, as all seven subunits were well resolved without any image processing. We also found that chemical fixation was necessary for the contact mode AFM to image GroEL/ES complexes, and in the AFM images. GroEL with GroES bound can be clearly distinguished from those without. The GroEL/ES complex was about 5 nm higher than GroEL alone, indicating a 2 nm upward movement of the apical domains of GroEL. Using a slightly larger probe force, unfixed GroEL could be dissected: the upper heptamer was removed to expose the contact surface of the two heptamers. These results clearly demonstrate the usefulness of cross-linking agents for the determination of molecular structures with the AFM. They also pave the way for using the AFM to study the structural basis for the function of GroE system and other molecular chaperones.     

Micropipette suction for measuring piconewton forces of adhesion and tether formation from neutrophil membranes.

A new method for measuring piconewton-scale forces that employs micropipette suction is presented here. Spherical cells or beads are used directly as force transducers, and forces as small as 10-20 pN can be imposed. When the transducer is stationary in the pipette, the force is simply the product of the suction pressure and the cross-sectional area of the pipette minus a small correction for the narrow gap that exists between the transducer and the pipette wall. When the transducer is moving along the pipette, the force on it is corrected by a factor that is proportional to the ratio of its velocity relative to its drag-free velocity. With this technique, the minimum force required to form a membrane tether from neutrophils is determined (45 pN), and the length of the microvilli on the surface of neutrophils is inferred. The strength of this technique is in its simplicity and its ability to measure forces between cells without requiring a separate theory or a calibration against an external standard and without requiring the use of a solid surface.     

拟夏孢锈属（Uredinopsis）新记录种

采到骨状拟夏孢锈（Ｕｒｅｄｉｎｏｐｓｉｓ ｏｓｓｉｆｏｒｍｉｓ Ｋａｍｅｉ）为中国新记录种。     

红毛五加叶的三萜皂甙

红毛五加（ＡｃａｎｔｈｏｐａｎａｘｇｉｒａｌｄｉｉＨａｒｍｓ）系五加科五加属植物,分布于河北、河南、四川、陕西、甘肃等省,有祛风湿、通关节、强筋骨、治痿痹等功效［１］。其化学成分未见报道。本文报道红毛五加叶的５个三萜皂甙（甙Ⅰ、Ⅱ、Ⅲ、Ⅳ和Ⅴ）的分离鉴定。它们均首次从该属植物发现,其结构如下：　　　　　　　　　　　　　Ｒ１　　　　　　　　　　　　　　Ｒ２　　　　　　　　Ⅰ　ｒｈａｍ－（１→２）－ａｒａ　　　　　Ｈ　　　　　　　Ⅱ　Ｈ　　　ｒｈａｍ－（１→４）－ｇｌｃ－（１→６）－ｇｌｃ　　　　　…     

水平回转对水稻幼苗叶细胞的影响

对在模拟微重力装置上回转１４ 天的水稻幼苗叶细胞进行了亚显微形态、电子探针和细胞酶化学研究。发现叶细胞质膜上Ｃａ２＋ －ＡＴＰ酶活性消失,膜内钙总量上升、膜外钙总量下降,细胞骨架变得疏松,细胞壁变薄并凹凸不平。叶绿体的基粒和线粒体的内嵴亦有部分变化。其变化机制,首先是细胞质膜上Ｃａ２＋ －ＡＴＰ酶活性消失,膜上钙泵停止工作,跨膜钙浓度差减小,膜内钙浓度上升,微管、微丝聚合受阻,细胞骨架疏松,分泌泡移动失去导向,从而导致细胞壁变薄等状态     

Supercritical carbon dioxide extraction of hydrocarbons from the microalgaBotryococcus braunii

Samples of the microalgaBotryococcus braunii were submitted to supercritical fluid extraction with carbon dioxide at 40 °C and pressures of 12.5, 20.0 and 30.0 MPa. The extraction yield and the fraction of the hydrocarbons in the extracts both increased with pressure and at 30 MPa these compounds were obtained rapidly. This behaviour is associated with the localization of the hydrocarbons outside the cell wall. In the extracts, which are fluid, golden and limpid, chlorophyll and phospholipids were not detected.Author for correspondence     

Regulation of fibronectin and laminin binding activity in cultured human lymphoblastic cell lines

The current study shows that a clonal derivative of the Jurkat cell line up-regulates both the avidity and density of the α 6/β1 receptor in response to phorbol 12-myristate 13-acetate (PMA). This derivative attaches to fibronectin and, to a lesser degree, laminin constitutively. Adhesion and spreading are dramatically up-regulated following treatment with PMA. The response on fibronectin peaks within 4 hours, is insensitive to cyclohexaminde, can be blocked by monoclonal antibodies (Mabs) to the β1 and α 5 subunits of the β1 family of integrins, and is not associated with increased expression of the α 5 or β1 epitopes at the cell surface. In contrast, the response on laminin is biphasic. The early phase parallels the response on fibronectin. The second phase peaks after 48–72 hours of treatment with PMA, is sensitive to cycloheximide, can be blocked by Mabs to the β1 and α 6 subunits, and is associated with increased expression of the α 6 epitope. Both the density independent and dependent responses to PMA in Jurkat cells are blocked by the protein kinase inhibitor staurosporine. The HSB-2, CEM, Molt-4, and HPB-ALL T-lymphoblastic cell lines also up-regulate attachment to fibronectin and laminin following treatment with PMA. All four lines constitutively attach to fibronectin and show rapid up-regulation of attachment following treatment with PMA. None of the lines attach to laminin prior to PMA treatment; however, specific adhesion developed after 4–120 hours of treatment. The most mature lines (Jurkat and HPB-ALL) up-regulated adhesion on laminin more rapidly than the less phenotypically mature lines (CEM, Molt-4, and HSB-2). In summary, clonal derivatives of the Jurkat cell line up-regulated attachment to laminin through protein kinase dependent increases in α /β1 receptor avidity and density. In addition, the expression of functional receptors for laminin is linked to developmental maturity in a series of T-lymphoblastic cell lines. © 1993 Wiley-Liss, Inc.