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61.
In the search for candidate genes for the tuberous sclerosis (TSC1) disease locus on chromosome 9q34, we have isolated an overlapping series of 22 plasmid and phage cDNA clones covering nearly 7 kb and with an open reading frame of 5070 bp encoding a protein of 1690 amino acids. The putative protein product is a member of the kinesin superfamily and is homologous to the mouse KIF1A and theCaenorhabditas elegansunc-104 genes. Both KIF1A and unc-104 function in the anterograde axonal transport of synaptic vesicles. The human homolog is therefore termed H-ATSV (axonal transporter of synaptic vesicles, HGMW-approved nomenclature ATSV) Screening of DNA from 107 tuberous sclerosis patients and 80 unaffected individuals with H-ATSV cDNA probes by pulsed-field gel electrophoresis/Southern blotting following digestion by rare-cutting methylation-sensitive restriction enzymes showed variant banding patterns in three patients with tuberous sclerosis. However, further analysis indicated that these variant fragments represent a rare polymorphism probably associated with methylation of clustered restriction sites. There is no evidence to support H-ATSV as a candidate gene for TSC1. 相似文献
62.
Large scale production of recombinant mouse and rat growth hormone by fed-batch GS-NSO cell cultures
Zhou W Bibila T Glazomitsky K Montalyo J Chan C Distefano D Munshi S Robinson D Buckland B Aunins J 《Cytotechnology》1996,22(1-3):239-250
Investigations of biological effects of prolonged elevation of growth hormone in animals such as mice and rats require large amounts of mouse and rat growth hormone (GH) materials. As an alternative to scarce and expensive pituitary derived materials, both mouse and rat GH were expressed in NSO murine myeloma cells transfected with a vector containing the glutamine synthetase (GS) gene and two copies of mouse or rat GH cDNA. For optimal expression, the mouse GH vector also contained sequences for targeting integration by homologous recombination. Fed-batch culture processes for such clones were developed using a serum-free, glutamine-free medium and scaled up to 250 L production scale reactors. Concentrated solutions of proteins, amino acids and glucose were fed periodically to extend cell growth and culture lifetime, which led to an increase in the maximum viable cell concentration to 3.5×109 cells/L and an up to 10 fold increase in final mouse and rat rGH titers in comparison with batch cultures. For successful scale up, similar culture environmental conditions were maintained at different scales, and specific issues in large scale reactors such as balancing oxygen supply and carbon dioxide removal, were addressed. Very similar cell growth and protein productivity were obtained in the fed-batch cultures at different scales and in different production runs. The final mouse and rat rGH titers were approximately 580 and 240 mg/L, respectively. During fed-batch cultures, the cell growth stage transition was accompanied by a change in cellular metabolism. The specific glucose consumption rate decreased significantly after the transition from the growth to stationary stage, while lactate was produced in the exponential growth stage and became consumed in the stationary stage. This was roughly coincident with the beginning of ammonia and glutamate accumulation at the entry of cells into the stationary stage as the result of a reduced glutamine consumption and periodic nutrient additions. 相似文献
63.
An efficient impedance method was developed for rapid evaluation of cosmetic preservatives. The method used decimal reduction time or D-value to assess preservative efficacies. The D-value, which was calculated from the plot of Log CFU ml–1 versus time by linear regression analysis, could be obtained within 48 h. Thus, the time required for the challenge test was reduced from 4–8 weeks with the standard procedures (eg US Pharmacopeia), to 2 days with the current method. A calibration curve (r=-0.95) was established by plotting the Log CFU ml–1 versus capacitance detection time (DT) of 108 samples. With the calibration, CFU can be estimated directly from the impedance test without plating. Two commercial biocides and several other chemicals were evaluated in a shampoo by the impedance procedure againstPseudomonas aeruginosa. The D-values obtained from the impedance test were not significantly different from those produced by the conventional plate count method. The technique was found to be particularly useful when screening a large number of compounds to find novel preservatives and synergistic preservative combinations. 相似文献
64.
65.
HAU3是寄主范围很广的放线菌噬菌体。Southern杂交实验表明,HAU3可以整合到吸水链霉菌应城变种10-22和变铅青链霉菌66的突变体ZX1的染色体中,形成溶原,其溶原菌自发释放HAU3,不受热激和紫外线照射的诱导。通过比较HAU3衍生噬粒pIJ8300的DNA酶切片段在加热前、后电泳带谱的区别,将HAU3的cos位点在pIJ8300的图谱上得到了定位。还利用Southern杂交的方法定位了HAU3与宿主形成溶原时附着位点(attP),并利用脉冲电泳技术定位了在变铅青链霉菌ZX7和吸水链霉菌应城变种10-22中形成溶原的附着位点(attB)。这些信息均有利于以HAU3为基础的载体的发展和优化。 相似文献
66.
用人重组肿瘤坏死因子-α(Tumornecrosisfactor-α,TNF-α)和人天然α干扰素(Interferon-α-,IFN-α)在人胚胎肺纤维母细胞(HEF)和Hep-2细胞系上对常见呼吸道病毒所致细胞病变抑制进行比较观察。病毒包括不同型别的腺病毒5株,单疱病毒Ⅰ型(HSV-I)1株,鼻病毒1株,仙台病毒1株,VSV1株。结果提示TNF-α和IFN-α均具有广谱抗病毒活性。TNF-α的抑毒作用能被TNF-α申抗和IFN-β单抗完全去除,被IFN-α单抗部分去除TNF-α的抗病毒效应。TNF--α中和试验的结果提示:TNF抗病毒活性仍为IFN-β诱生所介导。 相似文献
67.
激光诱变选育AC10菜用大青豆的研究报告 总被引:2,自引:1,他引:1
采用有性杂交和激光红宝石辐照交替进行,经10多年的研究、试验、选育出一个蛋白质含量、脂肪含量特高的菜用大青豆-AC10。AC10菜用大青豆,系早熟、适应性广、抗逆性强、耐迟播、产量高、效益好的品种。全生育期110天、80-85天采摘青毛豆、单产鲜毛豆700公斤以上,老豆单产160公斤左右。据农业部谷物测试中心分析,蛋白质含量48.32%,脂肪含量21.36%,合计69.68%。查新结果表明,为国内 相似文献
68.
利用大鼠肝脏线粒体为材料,以琥珀酸为底物,研究了不同浓度的丹参酮Ⅱ-A磺酸钠对线粒体态4、态3呼吸及呼吸控制率,线粒体跨膜电位,线粒体呼吸链复合体(Ⅱ+Ⅲ)电子传递及质子转移活性的影响。结果证明丹参酮ⅡA-磺酸钠是线粒体呼吸链复合体(Ⅱ+Ⅲ)的有效抑制剂。文中对丹参酮ⅡA-磺酸钠在心肌缺血再灌注过程中的保护作用的分子机理进行了讨论。 相似文献
69.
A. J. Soitamo G. Zhou A. K. Clarke G. Öquist E-M. Aro P. Gustafsson 《Plant molecular biology》1994,26(2):709-721
The unicellular cyanobacterium Synechococcus sp. PCC 7942 has three psbA genes encoding two different forms of the photosystem II reaction centre protein D1 (D1:1 and D1:2). The level of expression of these psbA genes and the synthesis of D1:1 and D1:2 are strongly regulated under varying light conditions. In order to better understand the regulatory mechanisms underlying these processes, we have constructed a strain of Synechococcus sp. PCC 7942 capable of over-producing psbA mRNA and D1 protein. In this study, we describe the over-expression of D1:1 using a tac-hybrid promoter in front of the psbAI gene in combination with lacI
Q
repressor system. Over-production of D1:1 was induced by growing cells for 12 h at 50 mol photons m-2 s-1 in the presence of 40 or 80 g/ml IPTG. The amount of psbAI mRNA and that of D1:1 protein in cells grown with IPTG was three times and two times higher, respectively. A higher concentration of IPTG (i.e., 150 g/ml) did not further increase the production of the psbAI message or D1:1. The over-production of D1:1 caused a decrease in the level of D1:2 synthesised, resulting in most PSII reaction centres containing D1:1. However, the over-production of D1:1 had no effect on the pigment composition (chlorophyll a or phycocyanin/number of cells) or the light-saturated rate of photosynthesis. This and the fact that the total amounts of D1 and D2 proteins were not affected by IPTG suggest that the number of PSII centres within the membranes remained unchanged. From these results, we conclude that expression of psbAI can be regulated by using the tac promoter and lacI
Q
system. However, the accumulation of D1:1 protein into the membrane is regulated by the number of PSII centres. 相似文献
70.
Repeated polyketide synthase modules involved in the biosynthesis of a heptaene macrolide by Streptomyces sp. FR-008 总被引:4,自引:2,他引:2
Zhihao Hu Kai Bao Xiufen Zhou Qi Zhou David A. Hopwood Tobias Kieser Zixln Deng 《Molecular microbiology》1994,14(1):163-172
Genes for biosynthesis of a Streptomyces sp. FR-008 heptaene macrolide antibiotic with antifungal and mosquito larvicidal activity were cloned in Escherichia coli using heterologous DNA probes. The cloned genes were implicated in heptaene biosynthiesis by gene replacement. The FR-008 antibiotic contains a 38-membered, poiyketide-derived macrolide ring. Southern hybridization using probes encoding domains of the type i modular erythromycin polyketide synthase (PKS) showed that the Streptomyces sp. FR-008 PKS gene cluster contains repeated sequences spanning c. 105 kb of contiguous DNA; assuming c. 5 kb for each PKS module, this is in striking agreement with the expectation for the 21-step condensation process required for synthesis of the FR-008 carbon chain. The methods developed for transformation and gene replacement in Streptomyces sp. FR-008 make it possible to genetically manipulate polyene macrolide production, and may later lead to the biosynthesis of novel polyene macrolides. 相似文献