保藏菌种的先进方法

目前保藏细菌的常用方法有:①根据不同菌种低温保藏,隔双周或隔月移种传代,②在很低的温度下。使微生物细胞快速冻结且保持完整,然后在真空下使水分升华达到干燥,即冻干保存。前种操作简     

细胞培养狂犬病毒悬液的制备

<正> 狂犬病毒属于弹状病毒科,狂犬固定毒是一个实验室毒株,其特点是致病性低,能在幼地鼠肾细胞(BHK-21C13)中迅速繁殖,该细胞主要用于狂犬病毒复制研究和生产兽用疫苗。 对于严重暴露的病人,在疫苗自动免疫力产生之前,在伤口周围注射抗血清作局部被动抗体使用,可以收到良好的效果。但是,由于过多的被动抗体会抑制自动抗体的产生,因此必须调整好疫苗与抗血清的关系。 抗狂犬病血清传统的生产方法,是用狂犬固定毒注入兔脑内,待发病取脑制备悬液,再用于超免马生产抗血清。这种抗血清     

噬菌体T7基因6.5和6.7的克隆及其缺失变种的构建

 用适当的限制性内切酶,将噬菌体T7基因6.5和6.7从整个噬菌体T7基因组中分离出来,插入到质粒pBR322中去,转化E.Coli HMS174,筛选出这两个基因的成功克隆。运用同样手段,从整个噬菌体T7基因组中分离出含有部分基因6编码序列,而不含基因6.5和6.7编码序列的T7DNA片段,插入到pBR322的衍生质粒中去,转化Ecoli C1757,再用含有基因6和基因7的双突变噬菌体T7去感染这一转化菌,通过同源交叉而得到缺失基因6.5和6.7的噬菌体T7缺失变种。这种噬菌体只能在载有噬菌体T7基因6.5和6.7,或者只载有基因6.7质粒的寄主中增殖。通过噬菌体结构蛋白电泳分析证明,这种噬菌体丢失了野生型菌体T7所具有的两条结构蛋白带。     

温度和生长素对猕猴桃冬季扦插生根的影响

猕猴桃新近在世界上的许多地区均有种植。虽然曾采用许多常用的育苗法,以满足对苗木需求的日益增加,但唯有幼苗嫁接法和扦插法较为实用。软枝扦插发根相当容易,而硬枝扦插则偶有发根困难的现象。有关硬枝扦插的报道甚少,且试验结果不尽一致。应用底温22°—24℃或高至27℃,被认为是一项良好的技术,但尚未探明不同温度     

Nitrate inhibition of nodulation can be overcome by the ethylene inhibitor aminoethoxyvinylglycine

Previously, we reported (a) a positive correlation between the nitrate concentrations in growth medium and ethylene evolved from uninoculated and inoculated alfalfa (Medicago sativa) roots and (b) a negative correlation between ethylene evolution and nodulation. Here, we report that the inhibitory effect of NO₃⁻ on nodulation of alfalfa can be eliminated by the ethylene inhibitor aminoethoxyvinylglycine (AVG). This effect was probably related to the strong inhibition (90%) of ethylene biosynthesis caused by AVG in these inoculated and NO₃⁻-treated roots. These results support our hypothesis that the inhibitory effect of NO₃⁻ is mediated through the phytohormone ethylene. A possible role of endogenous ethylene in the autoregulation of nodulation also is discussed. AVG at 10 micromolar significantly (P < 0.05) increased total nitrogenase activity (acetylene reduction) in 2.5 and 5 millimolar NO₃⁻-fed plants probably as a result of the very high stimulation of nodulation.     

Stromal low temperature compartment derived from the inner membrane of the chloroplast envelope

Leaf discs of four dicotyledonous species, when incubated at temperatures of 4 to 18°C (optimum at 12°C) for 30 or 60 minutes, responded by accumulations of membranes in the chloroplast stroma in the space between the inner membrane of the envelope and the thylakoids. The accumulated membranes, here referred to as the low temperature compartment, were frequently continuous with the envelope membrane and exhibited kinetics of formation consistent with a derivation from the envelope. Results were similar for expanding leaves of garden pea (Pisum sativum), soybean (Glycine max), spinach (Spinacia oleracea), and tobacco (Nicotiana tabacum). We suggest that the stromal low temperature compartment may be analogous to the compartment induced to form between the transitional endoplasmic reticulum and the Golgi apparatus at low temperatures. The findings provide evidence for the possibility of a vesicular transfer of membrane constituents between the inner membrane of the chloroplast envelope and the thylakoids of mature chloroplasts in expanding leaves.     

Wheat Inhibitors of Heterologous alpha-Amylases : Characterization of Major Components from the Monomeric Class

The four major components of the wheat monomeric α-amylase inhibitors (WMAI) from wheat, Triticum aestivum, endosperm have been isolated and characterized. Two of them, WMAI-1 and WMAI-2, are highly active against the α-amylase from the insect Tenebrio molitor and their N-terminal amino acid sequences indicate that they are closely related to each other (86% identical residues) and to the other members of the family (subunits of dimeric and tetrameric α-amylase inhibitors and trypsin inhibitors). WMAI-1, which is identical to the previously described 0.28 inhibitor, is encoded by a gene located in the short arm of chromosome 6D and WMAI-2 by a gene in the short arm of chromosome 6B. Components 3 and 4, which have blocked N-terminal residues, have identical internal amino acid sequences and are a separate class of proteins with respect to WMAI-1 and WMAI-2, although their amino acid composition and apparent molecular weights are quite similar. Their inhibitory activity versus α-amylases is either unstable during the purification process or due to contamination with other inhibitors.     

In vivo regulatory phosphorylation site in c(4)-leaf phosphoenolpyruvate carboxylase from maize and sorghum

Reversible seryl-phosphorylation contributes to the light/dark regulation of C₄-leaf phosphoenolpyruvate carboxylase (PEPC) activity in vivo. The specific regulatory residue that, upon in vitro phosphorylation by a maize-leaf protein-serine kinase(s), leads to an increase in catalytic activity and a decrease in malate-sensitivity of the target enzyme has been recently identified as Ser-15 in ³²P-phosphorylated/activated dark-form maize PEPC (J-A Jiao, R Chollet [1990] Arch Biochem Biophys 283: 300-305). In order to ascertain whether this N-terminal seryl residue is, indeed, the in vivo regulatory phosphorylation site, [³²P]phosphopeptides were isolated and purified from in vivo ³²P-labeled maize and sorghum leaf PEPC and subjected to automated Edman degradation analysis. The results show that purified light-form maize PEPC contains 14-fold more ³²P-radioactivity than the corresponding dark-form enzyme on an equal protein basis and, more notably, only a single N-terminal serine residue (Ser-15 in maize PEPC and its structural homolog, Ser-8, in the sorghum enzyme) was found to be ³²P-phosphorylated in the light or dark. These in vivo observations, combined with the results from our previous in vitro phosphorylation studies (J-A Jiao, R Chollet [1989] Arch Biochem Biophys 269: 526-535; [1990] Arch Biochem Biophys 283: 300-305), demonstrate that an N-terminal seryl residue in C₄ PEPC is, indeed, the regulatory site that undergoes light/dark changes in phosphorylation-status and, thus, plays a major, if not cardinal role in the light-induced changes in catalytic and regulatory properties of this cytoplasmic C₄-photosynthesis enzyme in vivo.