检索结果-南通市图书馆

作者：邵晓南京中医药大学

学位级别：硕士

目的:三七和人参的主要药效物质基础均为达玛烷型人参皂苷。这些皂苷按皂苷元结构主要分为原人参二醇型和原人参三醇型两类,人参总皂苷中原人参三醇型皂苷比例偏多,三七总皂苷中则以原人参二醇型偏多;此外,三七总皂苷中还含有三七特有... 详细信息

目的:三七和人参的主要药效物质基础均为达玛烷型人参皂苷。这些皂苷按皂苷元结构主要分为原人参二醇型和原人参三醇型两类,人参总皂苷中原人参三醇型皂苷比例偏多,三七总皂苷中则以原人参二醇型偏多;此外,三七总皂苷中还含有三七特有的原人参三醇型皂苷:三七皂苷R1。众所周知,三七和人参在传统用法上存在较大的差异。物质基础决定药效的差异,那么三七和人参的药效差异是否为三七总皂苷中特有的皂苷或组分群所决定的,值得深入探索。为此,本次研究将利用成分敲除技术敲除掉三七总皂苷中的特有皂苷三七皂苷R1或者改变原人参二醇型皂苷与原人参三醇型皂苷含量的比值,制备三七总皂苷knock-out提取物,为后续探讨三七皂苷R1以及原人参二醇型皂苷在三七药效发挥中的作用提供材料保障,同时也为这两种敲除方法的优缺点积累数据。方法:实验1:树脂联用制备高纯度三七总皂苷在D101大孔树脂制备粗三七总皂苷的基础上,采用阴阳离子交换树脂对粗三七总皂苷进一步纯化,得到高纯度的三七总皂苷,并通过紫外吸光度法和香草醛比色法评估其纯度。实验2:UPLC-CAD柱后补偿结合内参法定量分析三七总皂苷采用超高效液相-电喷雾检测器(UPLC-CAD)柱后补偿结合内参物,定量分析三七总皂苷中各个单体皂苷的含量,评价三七总皂苷的纯度并计算原人参二醇型皂苷与原人参三醇型皂苷含量的比值,Rbl与Rgl皂苷含量的比值。实验3:制备液相法敲除三七总皂苷中的三七皂苷R1称取三七总皂苷样品适量,10%甲醇溶解,根据要敲除的三七R1皂苷的成分确定液相洗脱程序,流动相为水和乙腈。依据保留时间收集目标成分,同时收集剔除目标成分的溶液,60℃以下减压回收,从而得到所需的样品。UPLC-CAD柱后补偿法进一步分析三七皂苷R1的敲除情况及敲除前后原人参二醇型皂苷和原人参三醇型皂苷含量比值的变化情况。实验4:免疫亲和柱层析法敲除三七总皂苷中的三七皂苷R1三七皂苷R1与BSA用高碘酸钠法偶联,形成三七皂苷R1-BSA抗原。用该抗原免疫家兔,得到三七皂苷R1的抗血清。抗血清经硫酸铵沉淀去除大部分杂质后,利用蛋白A材料制备成免疫亲和柱。最后,基于抗原-抗体间的相互作用,采用免疫亲和柱层析法敲除三七总皂苷中的三七皂苷R1。收集的样品经UPLC-CAD柱后补偿法检测,分析三七皂苷R1的敲除情况及敲除前后原人参二醇型皂苷和原人参三醇型皂苷含量比值的变化情况。实验5:大孔树脂柱层析法改变三七总皂苷中原人参二醇型皂苷与原人参三醇型皂苷含量的比值三七总皂苷溶于100ml左右纯水中,反复过已预处理的D101大孔吸附树脂柱3次。将乙醇分为30%和80%一组,40%和80%一组,50%和80%一组,即一组先用低浓度的乙醇流经树脂柱,并收集液体为洗脱液1,随后用高浓度乙醇流经树脂柱,收集液体为洗脱液2,将两次洗脱液分别经60℃烘干,得到样品。最终一共收集到六个洗脱液样品。得到样品之后,同样用UPLC-CAD柱后补偿法检测各三七皂苷单体的含量。通过比较不同洗脱条件下原人参二醇型皂苷和原人参三醇型皂苷含量的比值,寻找改变三七总皂苷中原人参二醇型皂苷与原人参三醇含量比值的合理方案。结果:实验1:树脂D101、D201和D113的联用表现出优异的纯化效果。根据香草醛比色法,高纯度三七总皂苷中皂苷的含量可达约107%。结果表明,树脂联合除杂是中性天然极性组分的一种可行的纯化方法。实验2:UPLC-CAD柱后补偿法可成功将高纯度三七总皂苷中的主要成分分离且峰型相对清晰。内参法一测多评各单体皂苷的含量总和高达92%左右,同时,原人参二醇型皂苷与原人参三醇型皂苷含量的比值约为0.54,Rb1/Rg1的比值为0.37。实验3:制备液相法能够从三七总皂苷中成功敲除三七皂苷R1,敲除后原人参二醇型皂苷和原人参三醇型皂苷含量的比值为0.70,敲除后Rb1/Rg1含量的比值为1.82。实验4:免疫亲和法可以部分敲除三七皂苷R1,敲除后原人参二醇型皂苷和原人参三醇型皂苷含量的比值为0.53,敲除后Rb1/Rg1含量的比值为0.33。实验5:比较六组不同洗脱条件下原人参二醇型皂苷和原人参三醇型皂苷含量的比值,其中30%乙醇洗脱物的PPD/PPT含量比值最小,为0.26,Rb1/Rg1为0.20。50%乙醇洗脱,随后80%乙醇洗脱物的PPD/PPT含量比值最大,为2.5,Rbl/Rgl为2.38。结论:1.阴阳离子交换树脂联用可以制备出高纯度的三七总皂苷,同时通过UPLC-CAD柱后补偿结合内参法可以成功定量分析高纯度三七总皂苷中各单体皂苷的含量。2.色谱敲除和抗体敲除两种敲除方法均可以将三七皂苷R1从三七总皂苷内敲除,但是色谱敲除和抗体敲除相比,色谱敲除的效率更高且操作简便,敲除三七皂苷R1更完全,基本不改变原人参二醇型皂苷和原人参三醇型皂苷含量比值,可重复性好。而抗体敲除

关键词：三七总皂苷三七皂苷R1 原人参二醇原人参三醇 knock-out UPLC-CAD 制备液相免疫亲和

来源：

同方学位论文库评论

在线全文

同方学位论文库

学校读者我要写书评

暂无评论

Mutation of cytotoxin-associated gene A affects expressions of antioxidant proteins of Helicobacter pylori

引用

World Journal of Gastroenterology 2009年第5期15卷 599-606页

作者： Zhi-Gang Huang Guang-Cai Duan Qing-Tang Fan Wei-Dong Zhang Chun-Hua Song Xue-Yong Huang Rong-Guang Zhang Department of Epidemiology College of Public HealthZhengzhou University Zhengzhou 450001 Henan ProvinceChina Department of Epidemiology School of Public Health Guangdong Medical College Dongguan 523808Guangdong Province China

AIM: To determine if disruption of the cagA gene of Helicobacter pylori (H pylori) has an effect on the expression of other proteins at proteome level. METHODS: Construction of a cagA knock out mutant Hp27 _△cagA (cagA -) via homologous recombination with the wild-type strain Hp27 (cagA+) as a recipient was performed. The method of sonication-urea-CHAPS-DTT was employed to extract bacterial proteins from both strains. Soluble proteins were analyzed by two-dimensional electrophoresis (2-DE). Images of 2-DE gels were digitalized and analyzed. Only spots that had a statistical signif icance in differential expression were selected and analyzed by matrix-assisted laser desorption/ionizationtime of flight mass spectrometry (MALDI-TOF-MS). Biological information was used to search protein database and identify the biological function of proteins. RESULTS: The proteome expressions between wild-type strain and isogenic mutant with the cagA gene knocked-out were compared. Five protein spots with high abundance in bacteria proteins of wild-type strains, down-regulated or absently expressed in bacteria proteins of mutants, were identified and analyzed. From a quantitative point of view, the identified proteins are related to the cagA gene and important antioxidant proteins of H pylori , including alkyl hydroperoxide reductase (Ahp), superoxide dismutase (SOD) and modulator of drug activity (Mda66), respectively, suggesting that cagA is important to maintain the normal activity of antioxidative stress and ensure H pylori persistent colonization in the host. CONCLUSION: cagA gene is relevant to the expressions of antioxidant proteins of H pylori, which may be a novel mechanism involved in H pylori cagA pathogenesis.

关键词： Helicobacter pylorr, Cytotoxin-associatedgene A knock-out Antioxidant protein Two-dimensional electrophoresis

来源：

维普期刊数据库评论

在线全文

维普期刊数据库

学校读者我要写书评

暂无评论

Targeted Mutagenesis in Zea mays Using TALENs and the CRISPR/Cas System

引用

Journal of Genetics and Genomics 2014年第2期41卷 63-68页

作者： Zhen Liang Kang Zhang Kunling Chen Caixia Gao State Key Laboratory of Plant Cell and Chromosome Engineering Institute of Genetics and Developmental BiologyChinese Academy of Sciences

Transcription activator-like effector nucleases （TALENs） and clustered regularly interspaced short palindromic repeats （CRISPR）/ CRISPR-associated （Cas） systems have emerged as powerful tools for genome editing in a variety of species. Here, we report, for the first time, targeted mutagenesis in Zea mays using TALENs and the CRISPR/Cas system. We designed five TALENs targeting 4 genes, namely ZmPDS, ZmlPKIA, ZmlPK, ZmMRP4, and obtained targeting efficiencies of up to 23.1% in protoplasts, and about 13.3% to 39.1% of the transgenic plants were somatic mutations. Also, we constructed two gRNAs targeting the ZmlPK gene in maize protoplasts, at frequencies of 16.4% and 19.1%, respectively. In addition, the CRISPR/Cas system induced targeted mutations in Z. mays protoplasts with efficiencies （13.1%） similar to those obtained with TALENs （9.1%）. Our results show that both TALENs and the CRISPR/Cas system can be used for genome modification in maize.

关键词： TAL-effector nucleases CRISPR/Cas system knock-out Zea mays

来源：

维普期刊数据库

同方期刊数据库评论

在线全文

学校读者我要写书评

暂无评论

p53-dependent and independent apoptosis induced by ionizing radiation in murine bone marrow cells

引用

Chinese Science Bulletin 1997年第2期42卷 155-159页

作者： Cui, YF Zhou, PK Lord, BI Hendry, JH BEIJING INST RADIAT MED BEIJING 100850PEOPLES R CHINA

APOPTOSIS or programmed cell death is a model of cell death, along with typical morphological and biochemical changes. The cell death, occurring during embryonic development, T and B cell maturation and endocrine-induced atrophy, is a physiological apoptosis which controls the amount of cells in the body. Apoptosis can also be induced by various factors, such as ionizing radiation and anti-tumour agents which damage DNA, and has been proposed to be a consequence of the induced expression of some tumour-related genes or increase of the proteins sta-

关键词： apoptosis p53 knock-out mouse bone marrow cells ionizing radiation.

来源：

维普期刊数据库

同方期刊数据库评论

在线全文

学校读者我要写书评

暂无评论

CB1 receptor activation on VgluT2-expressing glutamatergic neurons underlies Δ9-tetrahydrocannabinol(Δ9-THC)-induced aversive effects in mice

引用

中国药理学与毒理学杂志 2018年第9期32卷 715-716页

作者： HAN Xiao HE Yi BI Guo-hua ZHANG Hai-ying SONG Rui LI Jin XI Zheng-xiong Beijing Institute of Pharmacology and Toxicology Beijing 100850 China Molecular Targets and Medications Discovery Branch Intramural Research Program National Institute on Drug Abuse Baltimore MD 21224 USA

OBJECTIVE Cannabis can be rewarding or *** reward is believed to be mediated by activation of cannabinoid CB1 receptors(CB1 Rs) on GABAergic neurons that disinhibit dopaminergic neurons in the ventral tegmental area(VTA).However,little is known about the mechanisms underlying cannabis aversion in *** study aimed at dig the mechanisms underlying cannabis *** We first created CB1-floxed mice and then generated conditional CB1-knockout mice(VgluT2-CB1-/-) in glutamatergic neurons that express vesicular glutamate transporter 2(VgluT2).We then used immunohistochemistry and RNAscope in situ hybridization assays to examine whether CB1 Rs are expressed in VTA GABAergic neurons and glutamatergic *** also used Cre-dependent viral vector to express light-sensitive channelrhodopsin-2 into VTA glutamatergic ***,conditioned place preference and intracranial self-stimulation(ICSS) maintained by optical activation of VTA glutamatergic neurons were employed to evaluate the effects of Δ9-THC on brain reward *** CB1 Rs are found not only on VTA GABAergic neurons,but also on VTA glutamatergic neurons that express vesicular glutamate transporter 2(VgluT2).Photoactivation of VTA glutamatergic neurons produced robust intracranial self-stimulation(ICSS) behavior,which was dose-dependently blocked by DA receptor antagonists,but enhanced by *** contrast,Δ9-tetrahydrocannabinol(Δ9-THC),the major psychoactive component of cannabis,produced dose-dependent conditioned place aversion and a reduction in the above optical ICSS in VgluT2-cre control mice,but not in VgluT2-CB1-/-*** Activation of CB1 Rs in VgluT2-expressing glutamate neurons produces aversive effects that might explain why cannabinoid is not rewarding in rodents and might also account for individual differences in the hedonic effects of cannabis in humans.

关键词： cannabis cannabinoid receptor type 1 RNAscope conditioned knock-out intracrnialself-stimulation

来源：

维普期刊数据库

同方期刊数据库评论

在线全文

学校读者我要写书评

暂无评论

欢迎您,

建议与咨询留下您的常用邮箱和电话号码，以便我们向您反馈解决方案和替代方法

时间限定

文献类型

馆藏选择

核心期刊

语言

文献类型

帮助

文字说明：

检索规则说明：

检索范例：

分类表

所选分类

限定检索结果

文献类型

馆藏范围

日期分布

学科分类号

主题

机构

作者

语言

在线全文

在线全文

在线全文

在线全文

在线全文

请选择保存的检索档案：

请选择收藏分类：

通借通还

欢迎您,

建议与咨询 留下您的常用邮箱和电话号码，以便我们向您反馈解决方案和替代方法

时间限定

文献类型

馆藏选择

核心期刊

语言

文献类型

帮助

文字说明：

检索规则说明：

检索范例：

分类表

所选分类

限定检索结果

文献类型

馆藏范围

日期分布

学科分类号

主题

机构

作者

语言

在线全文

在线全文

在线全文

在线全文

在线全文

请选择保存的检索档案： 新增检索档案 确定 取消

请选择收藏分类： 新增自定义分类 确定 取消

通借通还

建议与咨询留下您的常用邮箱和电话号码，以便我们向您反馈解决方案和替代方法

请选择保存的检索档案：

请选择收藏分类：