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1.
本文采用脉冲激光荧光技术及常规的荧光分光光度计详细地研究了顺铂对DNA变性温度的影响,DNA氢键受损的碱基对数随温度变化,DNA的G—C碱基含量对顺铂作用效果以及顺铂浓度的影响,初步探讨了抗癌药物顺铂对DNA的结构影响.  相似文献   

2.
人神经tau蛋白与DNA相互作用   总被引:4,自引:0,他引:4  
通过凝胶阻滞实验表明,人类神经tau能够与不同来源的DNA(λDNA, 质粒DNA以及PCR产物)相结合,形成tau-DNA复合物.每分子神经tau大约与长度为6~10 bp的核苷酸片段结合.原子力显微镜直接证实了tau与线性质粒DNA相结合形成串珠样的结构.采用单克隆抗体Tau-1,进行免疫组织化学的实验显示,神经tau不但弥散地分布在SY5Y细胞系的细胞质内,同时也存在于细胞核中,并形成染色致密的斑点.以上结果提示:神经tau在细胞核中可能参与了某种重要的生物学功能.  相似文献   

3.
激光作用质粒DNA和小牛胸腺DNA的AFM研究   总被引:3,自引:0,他引:3  
激光作用质粒DNA和小牛胸腺DNA产生损伤效应,导致DNA结构变化,利用一种改进的试样制备过程和纳米显微镜--原子力显微镜(AFM)能够获得可重现的激光作用质粒DNA和小牛胸腺DNA的AFM图象,显示它们的特殊的表面结构。  相似文献   

4.
DNA单分子近场光学成像与荧光探测   总被引:3,自引:0,他引:3  
介绍了扫描近场光学(SNOM-Scanning Near-Field Optical Microscope)/原子力显微镜(AFM-Atomic Force Microscope)系统(SNO/AM)的工作原理。在AFM模式和SNOM模式下对DNA分子进行成像和荧光探测,得到了清晰的DNA单分子的形貌像和荧光像。由形貌圆像得到的DNA分子尺寸横向为20nm,高度为2nm,其中包含了探针形貌的影响。实验中采Tapping模式的AFM成像,样品经多次搜索扫描无明显损坏。AFM模式的分辨率优于1nm。SNOM模式下DNA分子形貌像和荧光像清晰,由近场荧光分布可以确定分子取向和浓度。用YOYO-1染料对λDNA分子进行染色和荧光探测。通过对DNA分子多个截面进行测量,分析染料 与DNA结合状态。  相似文献   

5.
原子力显微镜单分子力谱研究生物分子间相互作用   总被引:2,自引:0,他引:2  
原子力显微镜单分子力谱是近年来发展起来的能在单分子水平研究生物分子相互作用的新工具。本文综述了单分子力谱的测定原理、方法及其在研究蛋白.蛋白、蛋白-DNA相互作用,蛋白质去折叠和活细胞上配体/受体结合中的应用进展。  相似文献   

6.
激光对DNA作用机理的AFM研究   总被引:8,自引:0,他引:8  
激光作用质粒DNA和小牛胸腺DNA产生损伤效应,导致DNA结构变化,利用一种改进的试样制备过程和纳米显微镜--原子力显微镜(AFM)能够获得可重现的激光作用质粒DNA和小牛胸腺DNA的AFM图像,显示它们的特殊的表达结构,讨论了激光辐照导致DNA链断裂的作用机理。  相似文献   

7.
目的:评价注射用奥沙利铂冻干粉针制剂的安全性。方法:血管刺激性实验:主要观察家兔耳缘静脉注射注射用奥沙利铂后48 h和2周的耳局部血管变化;溶血性实验:观察本品在4h内有无溶血和聚集现象;全身过敏性实验:四组豚鼠隔日腹腔注射注射用奥沙利铂、5%卵蛋白或5%葡萄糖溶液,每大组随机分成二小组,第一组于首次注射后第14天,第二组于首次注射后第21天,静脉注射本品,观察30 min内动物的变化。结果:注射用奥沙利铂对家兔血管无刺激作用,家兔红细胞无溶血现象,豚鼠未见过敏反应。结论:实验结果表明注射用奥沙利铂安全可靠。  相似文献   

8.
用生物微量热技术及DNATm测量研究了手性不同的三种环方铂络合物与小牛胸腺DNA(200bp)作用中的特异性,发现R,R构型的与DNA作用最强,这与癌细胞的体外筛选结果相一致.而且通过HPLC及13C-NMR研究为环方铂络合物与DNA作用的分子机理分析找到了直接的证据.  相似文献   

9.
目的:研究奥沙利铂(L-OHP)联合化疗治疗晚期结肠癌的疗效.方法:选取60例晚期结肠癌患者,随机分为治疗组和对照组,每组各30例,对照组给予常规化疗治疗,治疗组给予L-OHP联合化疗治疗,观察比较两组的疗效和不良反应.结果:治疗组治疗总有效率(53.3%)显著高于对照组(36.7%)(P<0.05);治疗组中初治患者治疗有效率(63.2%)显著高于复治患者(36.3%)(P<0.05),也显著高于对照组中的初诊患者治疗有效率(38.9%)(P<0.05).对照组中初治患者治疗有效率(38.9%)和复治患者(30.0%)相当(P>0.05),复治患者的治疗有效率与对照组复治患者相当(P>0.05).治疗组白细胞减少和神经毒性的发生率要高于对照组(P<0.05),而治疗组出现口腔炎的比例低于对照组(P<0.05),恶心呕吐、腹泻、血小板减少、手足综合征和肾毒性的发生率均相当(P>0.05).结论:L-OHP联合化疗治疗晚期结肠癌初治患者具有较好的近期疗效和安全性,但其远期疗效和预后尚需进一步研究观察.  相似文献   

10.
利用彗星电泳检测出UVB、UVC短时间照射会使肿瘤细胞的DNA发生断裂,而长时间照射之后彗星电泳无法检测到碎片,推测可能是由于DNA分子交联的原因[1],国内外尚无定论.为了更直观的研究这种现象,提取了UVB,UVA照射后K562细胞的DNA,并调节到合适的浓度在原子力显微镜下观测.实验结果表明UVB对K562肿瘤细胞DNA损伤的影响呈现时间/剂量效应,较短时间照射主要产生DNA的链断裂,较长时间辐射则主要产生DNA链的交联.UVC对K562肿瘤细胞DNA的损伤大于UVB.UVC短时照射即可引起DNA的断裂和交联,较长时间辐射主要产生交联和一些断裂;长时间照射不但产生大量交联,同时有大量断裂产生,并发生凝缩和缠绕等结构破坏.  相似文献   

11.
Linear DNA, circular DNA, and circular DNA complexes with trivaline (TV), a synthetic oligopeptide, were imaged by atomic force microscopy (AFM) using mica as a conventional supporting substrate and modified highly ordered pyrolytic graphite (HOPG) as an alternative substrate. A method of modifying the HOPG surface was developed that enabled the adsorption of DNA and DNA–TV complexes onto this surface. On mica, both purified DNA and DNA–TV complexes were shown to undergo significant structural distortions: DNA molecules decrease in height and DNA–TV displays substantial changes in the shape of its circular compact structures. Use of the HOPG support helps preserve the structural integrity of the complexes and increase the measured height of DNA molecules up to 2 nm. AFM with the HOPG support was shown to efficiently reveal the particular points of the complexes where, according to known models of their organization, a great number of bent DNA fibers meet. These results provide additional information on DNA organization in its complexes with TV and are also of methodological interest, since the use of the modified HOPG may widen the possibilities of AFM in studying DNA and its complexes with various ligands.  相似文献   

12.
Atomic force microscopy was used to image single-stranded DNA (ssDNA) adsorbed on mica modified by Mg(2+), by 3-aminopropyltriethoxysilane or on modified highly oriented pyrolytic graphite (HOPG). ssDNA molecules on mica have compact structures with lumps, loops and super twisting, while on modified HOPG graphite ssDNA molecules adopt a conformation without secondary structures. We have shown that the immobilization of ssDNA under standard conditions on modified HOPG eliminates intramolecular base-pairing, thus this method could be important for studying certain processes involving ssDNA in more details.  相似文献   

13.
一种基于单分子纳米操纵的有序化测序策略   总被引:1,自引:0,他引:1  
尽管包括人类在内的许多生物物种的基因组测序工作已经完成,但由于现有测序技术的限制,大部分复杂基因组还存在很多大大小小的缺口.缺口的填补以及对其他重复序列区域的测序迫切需要全新的思路和技术.基于在DNA单分子定位切割和拾取方面的实验进展,提出了一种基于原子力显微镜纳米操纵技术的单分子有序化测序策略.计算机模拟的结果表明,这一方法和策略是可行的,有助于解决目前测序工作中所遇到的一些棘手问题.  相似文献   

14.
15.
The influence of mica surface on DNA/ethidium bromide interactions is investigated by atomic force microscopy (AFM). We describe the diffusion mechanism of a DNA molecule on a mica surface by using a simple analytical model. It appears that the DNA diffusion on a mica surface is limited by the surface friction due to the counterion correlations between the divalent counterions condensed on both mica and DNA surfaces. We also study the structural changes of linear DNA adsorbed on mica upon ethidium bromide binding by AFM. It turns out that linear DNA molecules adsorbed on a mica surface are unable to relieve the topological constraint upon ethidium bromide binding. In particular, strongly adsorbed molecules tend to be highly entangled, while loosely bound DNA molecules appear more extended with very few crossovers. Adsorbed DNA molecules cannot move freely on the surface because of the surface friction. Therefore, the topological constraint increases due to the ethidium bromide binding. Moreover, we show that ethidium bromide has a lower affinity for strongly bound molecules due to the topological constraint induced by the surface friction.  相似文献   

16.
DNA gyrase negatively supercoils DNA in a reaction coupled to the binding and hydrolysis of ATP. Limited supercoiling can be achieved in the presence of the non-hydrolysable ATP analogue, 5'-adenylyl beta,gamma-imidodiphosphate (ADPNP). In order to negatively supercoil DNA, gyrase must wrap a length of DNA around itself in a positive sense. In previous work, the effect of ADPNP on the gyrase-DNA interaction has been assessed but has produced conflicting results; the aim of this work was to resolve this conflict. We have probed the wrapping of DNA around gyrase in the presence and in the absence of ADPNP using direct observation by atomic force microscopy (AFM). We confirm that gyrase indeed generates a significant curvature in DNA in the absence of nucleotide and we show that the addition of ADPNP leads to a complete loss of wrap. These results have been corroborated using a DNA relaxation assay involving topoisomerase I. We have re-analysed previous hydroxyl-radical footprinting and crystallography data, and highlight the fact that the gyrase-DNA complex is surprisingly asymmetric in the absence of nucleotide but is symmetric in the presence of ADPNP. We suggest a revised model for the conformation of DNA bound to the enzyme that is fully consistent with these AFM data, in which a closed loop of DNA is stabilised by the enzyme in the absence of ADPNP and is lost in the presence of nucleotide.  相似文献   

17.
Conformational changes in individual carboxymethylcellulose (CMC) chains deposited on a highly oriented pyrolytic graphite (HOPG) surface were investigated by atomic force microscopy (AFM). A small amount of CMC solution with various salt concentrations was deposited onto the HOPG surface. The CMC molecular chains adsorbed onto the HOPG surface were clearly visualized using tapping-mode AFM under ambient conditions, as compared with those on a hydrophilic mica surface. Each CMC chain was distinguishable at the molecular level based on the vertical profiles of the AFM images, and probably aligned along the HOPG crystal lattice. Higher NaCl concentrations brought about dramatic conformational changes from aligned single chains to globular aggregates via the molecular network structure only on the HOPG surface through electrostatic screening of the CM groups. Although CMC is a water-soluble hydrophilic polyelectrolyte, some interaction, possibly due to a CH-pi bonding between the glucopyranosic axial plane of CMC and the aromatic rings of HOPG, is considered to be effective and dominant for the unique molecular attachment. These phenomena would imply the potential use of HOPG as a substrate for not only molecular imaging, but also for nano-scale morphological control of cellulosic polymers and other structural polysaccharides.  相似文献   

18.
Jiang Y  Marszalek PE 《The EMBO journal》2011,30(14):2881-2893
In spite of extensive research, the mechanism by which MutS initiates DNA mismatch repair (MMR) remains controversial. We use atomic force microscopy (AFM) to capture how MutS orchestrates the first step of E. coli MMR. AFM images captured two types of MutS/DNA complexes: single-site binding and loop binding. In most of the DNA loops imaged, two closely associated MutS dimers formed a tetrameric complex in which one of the MutS dimers was located at or near the mismatch. Surprisingly, in the presence of ATP, one MutS dimer remained at or near the mismatch site and the other, while maintaining contact with the first dimer, relocated on the DNA by reeling in DNA, thereby producing expanding DNA loops. Our results indicate that MutS tetramers composed of two non-equivalent MutS dimers drive E. coli MMR, and these new observations now reconcile the apparent contradictions of previous 'sliding' and 'bending/looping' models of interaction between mismatch and strand signal.  相似文献   

19.
Plasmid pBR322 DNA (0.5mg/mL) isolated from Escherichia coli HB101 was suspended in Tris-HCl-EDTA (1 mol/L - 0.1 mol/L, pH8.5); then a drop of the above solution was deposited on freshly cleaved mica substrate. After adsorption for about 1 min, the sample was stained with phosphotungstic acid. The residua] solution was removed with a piece of filter paper. Afterwards the sample was imaged with a home-made atomic force microscope (AFM) in air. The AFM images of pBR322 DNA with a molecular resolution have been obtained. These images show that pBR322 DNA exists in several different topological structures: (i) relaxed circular DNA with a different diameter; (ii) supercondensed DNA with different particle sizes; (iii) dimeric catenane connected by one relaxed circular molecule and another dose-compacted molecule which might be either supercoiled or intramolecular knotted form; (iv) oligomeric catenane with multiple irregular molecules in which DNA is interlocked into a complex oligomer; (v) possibly-existing  相似文献   

20.
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