P—选择素凝集素—EGF功能域的克隆、表达及其单克隆抗体制备

制备特异性抗人P 选择素的凝集素 表皮生长因子 (L EGF)功能域的单克隆抗体。利用特异引物 ,通过RT PCR从外周血血小板中扩增出人P 选择素的L EGF功能域基因 ,将其克隆至pET42b( )载体中 ,测序验证后转染大肠杆菌BL2 1,经诱导表达了C端融合 6×His的蛋白质。融合蛋白质经分离纯化后 ,免疫Balb/c小鼠 ,应用杂交瘤技术 ,通过间接ELISA筛选阳性克隆。获得 3株可稳定分泌抗L EGF功能域单抗的杂交瘤细胞株 (B10、F3和H5 )。其亚型分别为IgG2 、IgG1和IgG3;轻链均为κ型。所获的单抗对LPS刺激活化的人脐静脉内皮细胞均有特异性结合反应 ,并可在体外阻断经凝血酶激活的血小板与中性粒细胞间的粘附。表明所获的单抗可特异性识别结合天然P 选择素 ,具有体外抗活化血小板与中性粒细胞粘附的功能 ,为进一步应用此单抗进行P 选择素结构和功能及抗粘附治疗研究提供了实验基础。     

Next-generation sequencing to generate interactome datasets

Next-generation sequencing has not been applied to protein-protein interactome network mapping so far because the association between the members of each interacting pair would not be maintained in en masse sequencing. We describe a massively parallel interactome-mapping pipeline, Stitch-seq, that combines PCR stitching with next-generation sequencing and used it to generate a new human interactome dataset. Stitch-seq is applicable to various interaction assays and should help expand interactome network mapping.     

适于癌基因表达数据集的新特征提取标准NFEC及其分类新算法研究

癌基因表达数据集具有小样本、高维数之特点,一般的机器学习机难以对其有效分类。因此,通常需要采用某些特征提取度量标准来进行降维处理。可是常用的一些特征提取度量标准亦会导致分类效果欠佳之问题。依据微分容量控制学习机DCCM,提出了一个新的特征提取度量标准NFEC,然后依据NFEC和DCCM,提出了适于癌基因表达数据集的特征提取算法DCCFE。实验表明,新的度量NFEC和新的特征提取算法DCCFE较之现有方法对癌基因表达数据集分类时更为有效。本文的工作意义在于:(1)提出了一个新的更有意义的特征提取度量标准;(2)DCCM可以采用比核函数更为一般的一阶可微函数,因而提出的新的特征提取算法更具普遍应用意义。     

The roles of autophagy in development and stress responses in Arabidopsis thaliana

Autophagy is a dynamic process that involves the recycling process of the degradation of intracellular materials. Over the past decade, our molecular and physiological understanding of plant autophagy has greatly been increased. Most essential autophagic machineries are conserved from yeast to plants. The roles that autophagy-related genes (ATGs) family play in the lifecycle of the Arabidopsis are proved to be similar to that in mammal. Autophagy is activated during certain stages of development, senescence or in response to starvation, or environmental stress in Arabidopsis. In the progression of autophagy, ATGs act as central signaling regulators and could develop sophisticated mechanisms to survive when plants are suffering unfavorable environments. It will facilitate further understanding of the molecular mechanisms of autophagy in plant. In this review, we will discuss recent advances in our understanding of autophagy in Arabidopsis, areas of controversy, and highlight potential future directions in autophagy research.     

NF-κB、IκB、IKK在非小细胞肺癌中的表达及意义

目的检测非小细胞肺癌(non-small cell lung cancer,NSCLC)中NF-κB P65、p-IκBα(IκBα磷酸化)、p-IKKβ(IKKβ磷酸化)的表达情况及其与NSCLC临床特征的关系。方法采用免疫组化Elivision法检测NF-κB P65、p-IκBα、p-KKβ在56例NSCLC中表达情况,以20例癌旁组织作为对照。结果在NSCLC中NF-κB P65、p-IκBα、p-IKKβ的表达阳性率分别为83.9%(47/56)、55.7%(31/56)、69.6%(39/56),癌旁组织三者分别为20%(4/20)、25%(5/20)、30%(6/20),NF-κB P65、p-IκBα、p-IKKβ的表达与吸烟史、TNM分期、淋巴结转移相关,差异有统计学意义(P<0.05)。结论 NF-κB P65、p-IκBα、p-IKKβ高表达与NSCLC的发生、发展起着重要作用。     

松果体昼夜节律生物钟分子机制的研究进展

在各种非哺乳类脊椎动物中 ,松果体起着中枢昼夜节律振荡器的作用。近来 ,在鸟类松果体中相继发现了几种钟基因 ,如Per、Cry、Clock和Bmal等 ,其表达的时间变化规律与哺乳类视交叉上核 (SCN)的非常相似。钟的振荡由其自身调控反馈环路的转录和翻译组成 ,鸟类松果体和哺乳类SCN似乎具有共同的钟振荡基本分子构架 ;若干钟基因产物作为正向或负向调节子影响钟的振荡 ;昼夜性的控时机制同时也需要翻译后事件的参与。这些过程对钟振荡器的稳定性和 /或钟导引的光输入通路有着重要的调控作用     

辽宁省辽河水生态系统健康评价

2009年6-8月,在对辽宁省辽河铁岭段、沈阳段、盘锦段20个断面水文、水质、着生藻类、栖息地状况实地调查的基础上,采用主成分分析方法,进行指标的筛选与指标权重的确定,构建了该河流水生态系统健康评价指标体系和健康评价标准体系,并用改进的灰色关联度法对辽河6个断面的水生态系统健康状况进行了评价.结果表明：辽宁省辽河3个评价断面的水生态系统健康程度一般,2个评价断面分别为较差和极差,仅有1个评价断面达到了亚健康程度,说明辽河河流水生态系统生态退化严重,需要进一步加强辽河水系的生态恢复及水环境污染综合治理.     

Generating gene knockout rats by homologous recombination in embryonic stem cells

We describe here a detailed protocol for generating gene knockout rats by homologous recombination in embryonic stem (ES) cells. This protocol comprises the following procedures: derivation and expansion of rat ES cells, construction of gene-targeting vectors, generation of gene-targeted rat ES cells and, finally, production of gene-targeted rats. The major differences between this protocol and the classical mouse gene-targeting protocol include ES cell culture methods, drug selection scheme, colony picking and screening strategies. This ES cell-based gene-targeting technique allows sophisticated genetic modifications to be performed in the rat, as many laboratories have been doing in the mouse for the past two decades. Recently we used this protocol to generate Tp53 (also known as p53) gene knockout rats. The entire process requires ～1 year to complete, from derivation of ES cells to generation of knockout rats.