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101.
将去除信号肽的人肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)cDNA插入到带有原核增强子样序列Px的新型表达载体pBV320中,使TNF cDNA 5′端直接置于大肠杆菌trp启动子下游,采用37℃恒温培养,使TNF在大肠杆菌中获得了高效表达,表达活性达1.35(±0.17)×10~6U/L菌液。表达的TNF-α对L929细胞的毒性作用可被抗人肿瘤坏死因子-α的单克隆抗体所中和。表达菌裂解液作SDS-聚丙烯酰胺凝胶电泳,显示有一条分子量与TNF分子量吻合、约为17000道尔顿的蛋白带。利用DEAE-Sepharose阴离子交换层析及Sephacryl S-200凝胶过滤对上述重组人TNF-α进行纯化,获得电泳纯产品,比活性为1.48×10~6U/mg。 相似文献
102.
103.
Jorge E. Moreira Arthur R. Hand L. A. Håkan Borg Stellan Sandler Michael Welsh Nils Welsh Décio L. Eizirik 《Virchows Archiv. B, Cell pathology including molecular pathology》1991,60(1):337-344
We have previously described a preferential reduction in the secretory response to nutrient secretagogues in pancreatic mouse
islets maintained in culture after in vitro exposure to streptozotocin (SZ). This reduction was associated with an impaired
substrate metabolism at the mitochondrial level. To further clarify this issue, mouse pancreatic islets were exposed in vitro
to 2.2 mM SZ for 30 min. At 4 h after SZ treatment ultrastructural changes were apparent in the endoplasmic reticulum and
Golgi areas of the B-cells. However, 2 and 6 days following SZ exposure the B-cells appeared well preserved, except for a
marked decrease in the number of insulin-containing secretory granules. A morphometric analysis of the B-cells 6 days after
SZ exposure showed a normal B-cell size and a normal volume fraction of B-cell mitochondria. However, there was a decrease
in total islet size and a 13% decrease in the volume fraction of B-cells in the islets. These mouse islets exhibited a decreased
content of the mitochondrial DNA-encoded cytochrome b mRNA, as evaluated by dot-blot analysis. As a whole, the data obtained
indicate that SZ treatment does not induce a decrease in the number of mitochondria or long-lasting ultrastructural damage
to this organelle. However, there is a clear decrease in the cytochrome b mRNA, suggesting that SZ can induce damage to the
mitochondrial DNA. 相似文献
104.
Phorbol esters enhance attachment of NIH/3T3 cells to laminin and type IV collagen substrates 总被引:2,自引:0,他引:2
The effect of phorbol esters on the adhesive properties of NIH/3T3 mouse fibroblasts was investigated using plastic substrates precoated with the extracellular matrix proteins fibronectin, collagen, and laminin. Treatment with phorbol 12-myristate 13-acetate (PMA) enhanced NIH/3T3 cell attachment to laminin and type IV collagen substrates but had little or no effect on attachment to fibronectin and type I collagen substrates. The effect of PMA in enhancing cell attachment to laminin and type IV collagen substrates was dose dependent between 10(-9) and 10(-7) M. PMA was effective as early as 30 min; the effect reached a maximum at 2 h and decreased gradually. Phorbol 12, 13-dibenzoate and phorbol 12, 13-diacetate were effective but to a lesser extent and phorbol 12-myristate and phorbol 13-acetate showed little or no effect. These results suggest that PMA may enhance NIH/3T3 cell adhesion through effects on laminin and type IV collagen receptors. Retinoic acid, which itself requires at least 6 h to show an effect on attachment, did not have any effect on cell attachment in 2 h and, if anything, slightly inhibited PMA-enhanced cell attachment to laminin and type IV collagen substrates. 相似文献
105.
106.
Effect of Naturally Occurring nif Reiterations on Symbiotic Effectiveness in Rhizobium phaseoli 总被引:2,自引:0,他引:2
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David Romero Paul W. Singleton Lorenzo Segovia Enrique Morett B. Ben Bohlool Rafael Palacios Guillermo Dvila 《Applied microbiology》1988,54(3):848-850
Most naturally occurring strains of Rhizobium phaseoli possess reiteration of the nif genes. Three regions contain nitrogenase structural genes in strain CFN42. Two of these regions (a and b) have copies of nifH, nifD, and nifK, whereas the third region (c) contains only nifH. Strains containing mutations in either nif region a or nif region b had significantly diminished symbiotic effectiveness compared with the wild-type strain on the basis of nodule mass, total nitrogenase activity per plant, nitrogenase specific activity, total nitrogen in the shoot, and percentage of nitrogen. A strain containing mutations in both nif region a and nif region b was totally ineffective. These data indicate that both nif region a and nif region b are needed for full symbiotic effectiveness in R. phaseoli. 相似文献
107.
Ben F. Koop Michael M. Miyamoto Jennifer E. Embury Morris Goodman John Czelusniak Jerry L. Slightom 《Journal of molecular evolution》1986,24(1-2):94-102
Summary We have mapped and sequenced the globin gene and seven surrounding Alu repeat sequences in the orangutan globin gene cluster and have compared these and other orangutan sequences to orthologously related human sequences. Noncoding flanking and intron sequences, synonymous sites of , , and globin coding regions, and Alu sequences in human and orangutan diverge by 3.2%, 2.7%, and 3.7%, respectively. These values compare to 3.6% from DNA hybridizations and 3.4% from the globin gene region. If as suggested by fossil evidence and molecular clock calculations, human and orangutan lineages diverged about 10–15 MYA, the rate of noncoding DNA evolution in the two species is 1.0–1.5×10–9 substitutions per site per year. We found no evidence for either the addition or deletion of Alu sequences from the globin gene cluster nor is there any evidence for recent concerted evolution among the Alu sequences examined. Both phylogenetic and phenetic distance analyses suggest that Alu sequences within the and globin gene clusters arose close to the time of simian and prosimian primate divergence (about 50–60 MYA). We conclude that Alu sequences have been evolving at the rate typical of noncoding DNA for the majority of primate history.Presented at the FEBS Symposium on Genome Organization and Evolution, held in Crete, Greece, September 1–5, 1986 相似文献
108.
固氮蓝藻高光放氢突变种的筛选和放氢特点 总被引:1,自引:1,他引:0
作者以前报道过几种快速生长的固氮蓝藻在某种条件下能好气光放氢,其速度可以达到光合放氧速度的10—15%,但这种活性只有在不积累氢气的流动气相下或在短时间内发生。本文报道用亚硝基胍诱变所得到的Anabaena spp。Strain CA的高光放氢突变种——N9A和18A——的筛选和氢代谢特点。在达生长饱和光照以后,野生型的光放氢活性与光照强度的增加成正相关,而其吸氢活性则与之成负相关,显示高光照强度可能抑制吸氢酶的活性。无论在什么光强下,均测不到两个突变种的吸氢活性,暗示在突变种中,吸氢酶或有关系统受损伤。把细胞固相化在琼脂上,在密闭系统中,高光强下培养50个小时,两个突变种光释放和积累的氢分别为野生型的2倍(N9A)和6倍(18A),后者等于氢占气相(1%CO_2的空气)的1.8%。两个突变种在生长速度、叶绿素含量、乙炔还原活性以及光合放氧方面与野生型无明显不同。当以含50—100nM的镍离子的培养基培养时,野生型的好气净产氢活性完全丢失,其吸氢活性却增加约10倍。培养基中镍离子的存在,对两个突变种的高光放氢活性则毫无影响,而且在此情况下,仍测不出其吸氢活性。实验结果表明,这两个突变种系吸氢酶缺陷型突变种。 相似文献
109.
从自然感染的意大利麻痹病蜂(APis mellifera)头部,经二次差速离心与蔗糖梯度离心获得纯化的慢性蜜蜂麻痹病病毒(CBPV)。纯化的CBPV制备物感染正常蜜蜂,4天后出现典型的麻痹症状,接着死亡,平均死亡率分别为95%与100%。SDS-聚丙烯酰胺凝胶电泳分析,二次差速离心初步纯化的病毒制备物含有多条蛋白带,而蔗糖密度梯度纯化的病毒制备物仅含有单一的多肽带。5%、7.5%与10%SDS-聚丙烯酰胺凝胶电泳分析,均检测出一种病毒蛋白质,分子量大约为24,200道尔顿,而且不同凝胶浓度检测的蛋白质分子量相近。慢性蜜蜂麻痹病病毒核酸也用SDS-聚丙烯酰胺凝胶电泳分析,结果表明,凝胶中有5条带,对核酸酶敏感,证明该病毒含有5个单股RNA组分。对慢性蜜蜂麻痹病病毒的基因组结构进行了讨论。 相似文献
110.
应用双晶X光荧光光谱仪研究生理性和病理性植物组织中某些元素的分布特征已有报道。本研究工作中采用日本国东芝厂出品的AFV701型双晶X光荧光光谱仪,测定并比较了植物感染病毒前、后的叶片中,不同硫状态相对比例的改变。应用此仪器首先测定各种标准硫化物硫状态的化学位移数值,在确定标准图谱及可分性之后,再测定感染蚕豆萎蔫病毒(BBWV)、黄瓜花叶病毒(CMV)、芜菁花叶病毒(TuMV)的茄、苋色藜、烟草和萝卜等四种寄主植物的显症病叶中不同硫状态的相对比例。发现应用此技术能明显地区别出病态植物与健康植物中硫状态相对比例的变化差异。 相似文献