双歧杆菌质粒聚合酶基因对质粒载体稳定性的影响

目的探讨双歧杆菌天然质粒的聚合酶基因（Bifidobacterium plasmid polymerase,BPP）对人工构建的大肠埃希菌～双歧杆菌穿梭质粒载体（shuttle vector）在长双歧杆菌中稳定性的影响。方法首先电转化质粒pBADs—A和pBADs-BPP至长双歧杆菌,培养鉴定后,接种含质粒pBADs—A和pBADs-BPP的双歧杆菌于AMP^-和AMP^＋的MRS培养液中。厌氧培养后,将样品涂布于AMP^＋的MRS固体培养板上计数菌落数。结果相同条件下质粒pBADs·BPP组菌落数高于质粒pBADs—A组（P〈0．05）。结论BPP可以增加质粒载体的稳定性。     

弗氏志贺菌磷酸化蛋白的检测

目的:检测弗氏志贺菌全菌蛋白中被磷酸化修饰的蛋白。方法:制取弗氏2a志贺菌2457T野生株全菌磷酸化蛋白样品时加入磷酸化酶抑制剂,随后对样品进行双向电泳,以抗磷酸丝氨酸/苏氨酸/酪氨酸抗体为免疫探针,通过Western印迹找到被磷酸化的蛋白,并进行胶内酶解及MALDI-TOF质谱分析。结果与结论:共检测到13个磷酸化蛋白,其中9个为代谢途径中的酶。     

Heparan sulfate proteoglycans regulate autophagy in Drosophila

Heparan sulfate-modified proteoglycans (HSPGs) are important regulators of signaling and molecular recognition at the cell surface and in the extracellular space. Disruption of HSPG core proteins, HS-synthesis, or HS-degradation can have profound effects on growth, patterning, and cell survival. The Drosophila neuromuscular junction provides a tractable model for understanding the activities of HSPGs at a synapse that displays developmental and activity-dependent plasticity. Muscle cell-specific knockdown of HS biosynthesis disrupted the organization of a specialized postsynaptic membrane, the subsynaptic reticulum (SSR), and affected the number and morphology of mitochondria. We provide evidence that these changes result from a dysregulation of macroautophagy (hereafter referred to as autophagy). Cellular and molecular markers of autophagy are all consistent with an increase in the levels of autophagy in the absence of normal HS-chain biosynthesis and modification. HS production is also required for normal levels of autophagy in the fat body, the central energy storage and nutritional sensing organ in Drosophila. Genetic mosaic analysis indicates that HS-dependent regulation of autophagy occurs non-cell autonomously, consistent with HSPGs influencing this cellular process via signaling in the extracellular space. These findings demonstrate that HS biosynthesis has important regulatory effects on autophagy and that autophagy is critical for normal assembly of postsynaptic membrane specializations.     

多甲氧基黄酮对人胶质母细胞瘤细胞系生物学特性的影响

目的：多甲氧基黄酮（PMFs）因其抗氧化、抗癌、抗炎及抗动脉粥样硬化等多样的生物学活性受到国内外学者的广泛关注,但其在神经胶质瘤治疗中的潜在作用尚未见报道。本研究对多甲氧基黄酮处理人胶质母细胞瘤细胞系对其生物学特性的影响,初步探讨PMFs在神经胶质瘤治疗中的潜在应用。方法：不同浓度（0,20,40,60,80,100μg／mL）的PMFs处理人胶质母细胞瘤细胞系U251不同时间后．分别用Annexinv／PI双染法检测细胞凋亡的变化,MTT法检细胞活力的变化,Transwell小室实验检测细胞侵袭能力的变化。结果：随着多甲氧基黄酮的浓度及处理时间的增加,细胞的增殖明显受到抑制,同时诱导细胞大量凋亡,促使细胞生长停滞于G2／M期;此外,多甲氧基黄酮剂量依赖性的抑制胶质瘤细胞的侵袭。结论：PMFs能剂量和时间依赖性降低人胶质母细胞瘤细胞系U251的,同时显著诱导细胞瘤的凋亡和侵袭能力,提示PMFs可能对神经胶质瘤的高增殖性和侵袭性有一定的抑制作用,因此可能具有成为治疗神经胶质瘤药物的潜在应用价值。     

灵芝两个无孢品种中水提物成分分析及免疫活性比较

龙芝2号和鹿角灵芝均为赤芝的栽培品种,且在生长发育过程中均不产生孢子。目前,尚缺乏对两者化学成分和药理活性的系统比较研究。本研究以灵芝两个无孢品种——龙芝2号和鹿角灵芝为原料,研究对比两者子实体水提物成分差异及免疫活性的强弱。采用化学及仪器分析相结合的方法,分析两者水提物中的多糖得率、含量、重均分子量分布特征及核苷、蛋白质、氨基酸含量差异,并研究了灵芝两个无孢品种水提物样品刺激RAW 264.7细胞释放NO活性。结果表明,两者多糖得率及含量差异不大,但龙芝2号中重均分子量分布范围较广,多糖分子量较大,其含有3种多糖,分子量分别为2.021×10⁶、1.802×10⁶和4.825×10⁵,而鹿角灵芝中仅含有1种多糖,分子量为1.589×10⁴;两者中含有的核苷种类相似,但各核苷的含量存在差异;鹿角灵芝和龙芝2号中蛋白质含量分别为10.70%和10.32%,两者均不含有组氨酸,蛋氨酸在两者中含量均较高,分别达到2.556%和2.591%。从鹿角灵芝和龙芝2号中得到的水提物样品均具有体外刺激巨噬细胞释放NO的活性。     

突触泡蛋白2研究进展

突触泡蛋白2(SV2)是一类跨膜糖蛋白,定位于脊椎动物神经元及内分泌细胞,与神经递质的释放、内分泌泡胞吐作用、突触泡稳态的维持、神经肌肉接头的形成及肾上腺素能受体α2C的定位密切相关。最近还发现SV2是肉毒神经毒素BoNT/A的受体,介导BoNT/A进入神经元。SV2可作为突触泡标记蛋白,广泛应用于生物学研究及肿瘤诊断。此外,SV2还是抗癫痫药物的作用靶标。     

Modified heparin inhibits P-selectin-mediated cell adhesion of human colon carcinoma cells to immobilized platelets under dynamic flow conditions

Accumulating evidence indicates that the formation of tumor cell platelet emboli complexes in the blood stream is a very important step during metastases and that the anti-metastasis effects of heparin are partially due to a blockade of P-selectin on platelets. In this study, heparin and chemically modified heparins were tested as inhibitors of three human colon carcinoma cell lines (COLO320, LS174T, and CW-2) binding to P-selectin, adhering to CHO cells expressing a transfected human P-selectin cDNA, and adhering to surface-anchored platelets expressing P-selectin under static and flow conditions. The aim was to screen for heparin derivatives with high anti-adhesion activity but negligible anticoagulant activity. In this study, four modified heparins with high anti-adhesion activity were identified including RO-heparin, CR-heparin, 2/3ODS-heparin, and N/2/3DS-heparin. NMR analysis proved the reliability of structure of the four modified heparins. Our findings suggested that the 6-O-sulfate group of glucosamine units in heparin is critical for the inhibition of P-selectin-mediated tumor cell adhesion. Heparan sulfate-like proteoglycans on these tumor cell surfaces are implicated in adhesion of the tumor cells to P-selectin. Some chemically modified heparins with low anticoagulant activities, such as 2/3ODS-heparin, may have potential value as therapeutic agents that block P-selectin-mediated cell adhesion and prevent tumor metastasis.     

草食动物对结缕草种子散布及发芽力的影响

研究了草食动物(牛、山羊)对辽东半岛结缕草种子散布及发芽力的影响。结果表明:风干牛粪、羊粪中结缕草种子的数量分别为101.7和6.88粒.g-1;牛、羊的采食和消化对结缕草种子的生活力没有影响,但提高了其发芽率(分别为74.12%和67.77%)、发芽势(分别为24.34%和17.72%)及活力指数(分别为41.86和37.38),缩短了其发芽时间(分别为12.53和12.70d);牛粪和羊粪中结缕草种子的发芽率虽有提高但没有达到其生活力水平;牛对结缕草种子散布及发芽力的影响效果均好于山羊。     

植物激素脱落酸受体的研究进展

脱落酸(abscisic acid, ABA)广泛参与植物生长发育的调控和对多种环境胁迫的适应性反应。有关ABA受体的研究已经在检测受体位置、纯化ABA特异性的结合蛋白和克隆ABA受体基因方面做出了许多重要的工作。最近相继发现一种RNA结合蛋白FCA和一种编码Mg离子螯合酶（Mg-chelatase）H亚基的CHLH作为两种不同的ABA受体分别调控植物的开花时间和介导种子萌发、幼苗生长及叶片的气孔运动。本文从实验策略的角度重点分析总结了研究脱落酸受体相对有效的途径与方法, 同时就有关的研究结果给予了评论和展望。     

不同粪便DNA提取方法比较分析北大核心CSCD

肠道微生物群落结构和多样性与人体疾病密切相关。然而,相关群落结构分析结果可能受到DNA提取质量等实验因素影响。因此,评估不同DNA提取方法对肠道特定种属的提取效果,对于全面、准确获取人体肠道微生物谱,深入探究肠道微生物群落结构具有指导意义。本研究旨在借助实时荧光定量PCR(real-time quantitative polymerase chain reaction,RT qPCR)技术,以DNA提取纯度、浓度,以及对肠道中特定种属微生物基因组DNA的提取丰度为指标,对5种DNA提取方法进行比较分析。结果表明,试剂盒Q的提取效果最佳,特别是对乳杆菌属和双歧杆菌属等革兰氏阳性菌的提取效果较好。N试剂盒的平均DNA提取浓度较Q试剂盒低,但在纯度方面,二者无显著性差异。与其他3种商用试剂盒(M、PSP、TG)相比,N方法对肠道内指定微生物基因组的提取效果仅次于Q试剂盒,位居第二。相比之下,M试剂盒提取所得DNA,质量较高,但浓度偏低,对于肠道内革兰氏阳性菌的提取效果不很理想。TG试剂盒和PSP试剂盒提取所得DNA在浓度、质量以及细菌丰度方面均不及其他验证的试剂盒。综上,Q试剂盒可作为肠道微生态研究相关实验中获取高质量基因组DNA的提取方法。本研究结果为肠道微生态研究相关实验中基因组DNA提取方法的选择提供参考依据。