中性红荧光探针法测定生物大分子核酸

中性红 (NR)是一种吩嗪染料 ,至今已有许多关于 NR与 DNA相互作用的报道[1～ 5] .李克安[4 ] 和黄承志等 [5]利用共振光散射技术分别在酸性 (p H=2 .3 )和中性 (p H=7.6～ 7.8)条件下 ,建立了以 NR为探针测定痕量 DNA的方法 .我们 [2 ,3]曾利用荧光光谱方法研究了在 p H=7.4条件下 NR与 DNA之间的相互作用 ,发现利用吖啶橙和 NR之间的能量转移现象可以测定 DNA,但检出限偏高 ,且由于使用两种染料试剂 ,操作较繁琐 .为了克服吖啶橙、NR能量转移分析法的不足 ,本文建立了在 p H=4.5的条件下以单一染料 NR为荧光探针测定痕量核酸的…     

Simple and Sensitive Assay for Nucleic Acids Using their Quenching Effect on the Chemiluminescence Reaction Between Luminol and Hydrogen Peroxide with Manganese-Tetrasulfonatophthalocyanine as a New Catalyst

The manganese-tetrasulfonatophthalo-cyanine (MnTSPc) catalyzed luminol-hydrogen peroxide chemiluminescence (CL) system can be quenched in the presence of nucleic acids. A new and highly sensitive CL quenching method for determining nucleic acids in aqueous solutions has been developed. Under optimum conditions, linear relationships were found between the quenched intensity of CL and the concentration of nucleic acids in the range 0.10–2.0µgmL^–1 for calf thymus DNA and 0–1.6µgmL^–1 for fish sperm DNA. The limits of detection were 14.8ngmL^–1 for calf thymus DNA and 21.7ngmL^–1 for fish sperm DNA. The relative standard deviation (RSD) of nine replicate measurements is 1.4% for 1.0µgmL^–1 calf thymus DNA. The method was applied to the analysis of nucleic acids in synthetic samples and the results are satisfactory.Received December 2, 2002; accepted June 2, 2003 published online September 1, 2003     

Determination of Nucleic Acids Using Rivanol as the Fluorescent Probe in the Presence of SDS

ABSTRACT

The quantitative method for nucleic acids using rivanol as the fluorescent probe in the presence of SDS was proposed. Under proper conditions, addition of nucleic acids to a mixture of rivanol and SDS resulted in enhanced fluorescence and spectral shifts of rivanol-SDS system. The calibration graph was linear in the range of 0–10.0 μg/ml for CT DNA and 0–9.0 μg/ml for yeast RNA, the limit of detection was 62 ng/ml for CT DNA and 156 ng/ml for yeast RNA. CT DNA could be determined in the presence of 20%(w/w) yeast RNA.     

喹哪啶蓝与核酸作用的光谱研究及分析

外部堆积结合;分光光度法;喹哪啶蓝与核酸作用的光谱研究及分析     

醋酸氯己定与甲酚红的荷移反应及应用

研究并确定了醋酸氯已定与甲酚红的荷移反应条件。在乙醇-丙酮介质中形成1：1型络合物,其λmax=455nm,醋酸氯已定浓度在0～80mg/L范围内呈线性关系;表观摩尔吸光系数为1．08×104L·mol-1·cm-1;桑德尔灵敏度是0．0345μg·cm-2;相对标准偏差小于1％（n＝8）,回收率符合要求。     

Studying the mechanism of RNA separations using RNA chromatography and its application in the analysis of ribosomal RNA and RNA:RNA interactions

DNA/RNA chromatography presents a versatile platform for the analysis of nucleic acids. Although the mechanism of separation of double stranded (ds) DNA fragments is largely understood, the mechanism by which RNA is separated appears more complicated. To further understand the separation mechanisms of RNA using ion pair reverse phase liquid chromatography, we have analysed a number of dsRNA and single stranded (ss) RNA fragments. The high-resolution separation of dsRNA was observed, in a similar manner to dsDNA under non-denaturing conditions. Moreover, the high-resolution separation of ssRNA was observed at high temperatures (75 °C) in contrast to ssDNA. It is proposed that the presence of duplex regions/secondary structures within the RNA remain at such temperatures, resulting in high-resolution RNA separations. The retention time of the nucleic acids reflects the relative hydrophobicity, through contributions of the nucleic sequence and the degree of secondary structure present. In addition, the analysis of RNA using such approaches was extended to enable the discrimination of bacterial 16S rRNA fragments and as an aid to conformational analysis of RNA. RNA:RNA interactions of the human telomerase RNA component (hTR) were analysed in conjunction with the incorporation of Mg²⁺ during chromatography. This novel chromatographic procedure permits analysis of the temperature dependent formation of dimeric RNA species.     

Study of the Reaction between Nucleic Acids and Tb-BPMPHD-CTMAB Complex and its Analytical Application

The direct use of the fluorescence emission properties of nucleic acids in the investigation of their biological properties is limited, whereas the use of metal complexes as fluorescence probe to study nucleic acids has increased remarkbly. Recently we found that the Tb-l,6-bi(1'-phenyl-3'-methyl-5'-pyrazolone-4'-)hezane-dione(BPMPHD)-trimethyl am-monium bromide(CTMAB) complex can be used as a sensitive fluorescence probe for the determination of nucleic acids.     

The Unexpected Base-Pairing Behavior of Cyanuric Acid in RNA and Ribose versus Cyanuric Acid Induced Helicene Assembly of Nucleic Acids: Implications for the Pre-RNA Paradigm

The cyanuric acid (CA) heterocycle forms supramolecular structures with adenine nucleobases/nucleosides and oligonucleotides, leading to speculation that they can act as forerunners to RNA. Herein, the assembly behavior of RNA containing CA and CA–ribose nucleoside was studied. Contrary to previous reports, CA in RNA and the CA-ribonucleoside resulted in destabilization of supramolecular assemblies, which led to a reevaluation of the CA–adenine hexameric rosette structure. An unprecedented noncovalent supramolecular helicene structure is proposed to account for the striking difference in behavior, which has implications for novel paradigms for reorganizing the structures of nucleic acids, the synthesis of long helicenes, and pre-RNA world paradigms. The results caution against extrapolating the self-assembly behavior of individual heterocycles from the level of monomers to oligomers because the base-paring properties of (non-)canonical nucleobases are impacted by the type of oligomeric backbone to which they are attached.     

DNA银纳米簇在功能核酸荧光生物传感器中的应用

DNA银纳米簇(DNA-AgNCs)是以DNA为模板, 通过碱基杂环上的N原子与Ag⁺结合, 用NaBH₄将Ag⁺还原得到的具有荧光性质的新兴纳米探针. 由于DNA-AgNCs具有合成方法简单、 生物相容性好和荧光发射波长可调等优点, 使其在分析检测等领域具有广泛的应用. 本文对DNA-AgNCs的合成和荧光性质两个方面进行了综述, 分类总结了以DNA-AgNCs为无标记荧光探针在功能核酸荧光生物传感器方面的应用, 对其不足与应用潜力进行展望, 以期为未来的研究与应用提供借鉴.     

DNA and RNA binding properties of an arginine-based ‘Extended Chiral Box’ Peptide Nucleic Acid

Lys-based ‘chiral box’ Peptide Nucleic Acids (PNAs with three adjacent 2D-Lys-based chiral monomers) have shown unsurpassed specificity in DNA recognition. In this Letter, the binding performances of arginine-based chiral PNAs were evaluated for PNAs containing in the middle part of the strand either a 2D,5L-Arg monomer or three adjacent 2D-; 2D,5L-; 5L-Arg monomers (‘Extended Chiral Box’), a combination never studied before. The binding performances of the PNAs were studied by evaluating the melting temperatures of fullmatch and mismatch PNA-DNA and PNA-RNA hybrids and by studying their structure by circular dichroism (CD). The data indicated that the arginine side chains inserted in the PNA structure are perfectly equivalent to lysine side chains as far as oligonucleotide recognition is concerned. The insertion of an ‘Extended Chiral Box’ into PNA differently influences the binding properties to DNA and RNA: the additional side chains had no observable effect on binding affinity and selectivity toward DNA, whereas, seemed to slightly disturb the binding affinity to RNA but at the same time highly enhancing the recognition selectivity.     

Efficient Oligomerization of Aromatic Amino Acids Induced by Gaps in Four-Helix Bundles of DNA or RNA

The formation of peptides from amino acids is one of the processes associated with life. Because of the dominant role of translation in extant biology, peptide-forming processes that are RNA induced are of particular interest. We have previously reported the formation of phosphoramidate-linked peptido RNAs as the products of spontaneous condensation reactions between ribonucleotides and free amino acids in aqueous solution. We now asked whether four-helix bundle (4HB) DNA or RNA folding motifs with a single- or double-nucleotide gap next to a 5’-phosphate can act as reaction sites for phosphoramidate formation. For glycine, this was found to be the case, whereas phenylalanine and tryptophan showed accelerated formation of peptides without a covalent link to the nucleic acid. Free peptides with up to 11 tryptophan or phenylalanine residues were found in precipitates forming in the presence of gap-containing DNA or RNA 4HBs. Control experiments using motifs with just a nick or primer alone did not have the same effect. Because folded structures with a gap in a double helix are likely products of hybridization of strands formed in statistically controlled oligomerization reactions, our results are interesting in the context of prebiotic scenarios. Independent of a putative role in evolution, our findings suggest that for some aromatic amino acids an RNA-induced pathway for oligomerization exists that does not have a discernable link to translation.     

溴连苯三酚红体系催化光度法测定亚硝酸根的研究

基于亚硝酸根以溶解氧氧化溴连苯三酚红的催经作用，建立了双波长分光光度动力学测定亚硝酸根的新方法，于３９２ｎｍ处测量体系吸光度Ａ１，４８４ｎｍ处测量体系吸光度Ａ２，以△Ａ＝Ａ１－Ａ２进行亚硝酸根定量，所建立的方法对亚硝酸根的线性范围为０．００５～１．０μｇ／ｍＬ。该法简便，灵敏，选择性好，应用于蔬菜样品的测定，结果满意。     

Spectroscopic Study of the Interaction between New Fluorescent Dyes with Nucleic Acids

Considerableefforthasbeencofltinuingtofocusonthedevelopmentofnewfluorescentdyestorecognizenucleicacids'-'.Althoughdansylisawell-knownsensitivehydrophobicprobewhichhasbeenwidelyutilizedasafluorescentprobeforthestudyofproteins,yetlittleefforthasbeenfocusedontheexploringdansylamidederivativeswhichmayhavespecificeffectsonnucleicacids.Sincethebindingaffinityofsuchfluorophorestopolynucleotideswasgreatlyaffectedbytheirsidechainsubstitutions,inthisworkseveralnewdansylderivativeswithspecificbindingtonu…     

更生霉素-DNA反应的电化学研究

研究了更生霉素(Act-D)在活化浸蜡石墨电极上的电化学特性,发现在-0.19和-0.51V分别出现一个灵敏的氧化峰和还原峰,加入DNA使Act-D的峰电流明显降低,生成了非电活性络合物.采用电化学方法研究了Act-D与小牛胸腺DNA的结合平衡,根据不可逆电活性分子与DNA相互作用的电化学公式对实验数据进行非线性拟合,获得了Act-D的扩散系数、Act-D与DNA的结合常数和结合位点数等重要信息,发现Act-D与DNA的8个碱基对紧密结合,其侧链加强了结合程度.     

Electron attachment to DNA and RNA nucleobases: An EOMCC investigation

We report a benchmark theoretical investigation of both vertical and adiabatic electron affinities of DNA and RNA nucleobases: adenine, guanine, cytosine, thymine, and uracil using equation of motion coupled cluster method. The vertical electron affinity (VEA) values of the first five states of the DNA and RNA nucleobases are computed. It is observed that the first electron attached state is energetically accessible in gas phase. Furthermore, an analysis of the natural orbitals exhibits that the first electron attached states of uracil and thymine are valence‐bound in nature and undergo significant structural changes on attachment of an extra electron, which reflects in the deviation of the adiabatic electron affinity (AEA) than that of the vertical ones. Conversely, the first electron attached states of cytosine, adenine, and guanine are in the category of dipole‐bound anions. Their structure, by and large, remain unaffected on attachment of an extra electron, which is evident from the observed small difference between the AEA and VEA values. VEA and AEA values of all the DNA and RNA nucleobases are found to be negative, which implies that the first electron attached states are not stable rather quasi bound. The results of all previous theoretical calculations are out of track and shows large deviation with respect to the experimentally measured values, whereas, our results are found to be in good agreement. Therefore, our computed values can be used as a reliable standard to calibrate new theoretical methods. © 2015 Wiley Periodicals, Inc.     

奈替米星-刚果红分光光度法测定奈替米星

在pH4.5~6.0和pH1.9~3.1的酸性条件下,奈替米星(NTM)与刚果红(CR)反应,分别生成红色和蓝色离子缔合物,使刚果红红色溶液褪色。前者最大褪色波长位于500nm处,表观摩尔吸光系数(ε)为2.55×104Lmol-1cm-1;奈替米星在0.04~9.5mg/L浓度范围内,遵从朗伯比尔定律。后果者也使奈替米星在一定浓度范围内遵从朗伯比尔定律,但灵敏度不及前者。本文探讨了适宜的反应条件、主要分析化学性质及两种不同酸度范围内的络合比。所拟定的方法简便、快速,用于市售奈替米星药物及人尿、人血清中奈替米星含量的测定,结果满意。     

RNA–DNA Chimeras in the Context of an RNA World Transition to an RNA/DNA World

The RNA world hypothesis posits that DNA and proteins were later inventions of early life, or the chemistry that gave rise to life. Most scenarios put forth for the emergence of DNA assume a clean separation of RNA and DNA polymer, and a smooth transition between RNA and DNA. However, based on the reality of “clutter” and lack of sophisticated separation/discrimination mechanisms in a protobiological (and/or prebiological) world, heterogeneous RNA–DNA backbone containing chimeric sequences could have been common—and have not been fully considered in models transitioning from an RNA world to an RNA–DNA world. Herein we show that there is a significant decrease in Watson–Crick duplex stability of the heterogeneous backbone chimeric duplexes that would impede base‐pair mediated interactions (and functions). These results point to the difficulties for the transition from one homogeneous system (RNA) to another (RNA/DNA) in an RNA world with a heterogeneous mixture of ribo‐ and deoxyribonucleotides and sequences, while suggesting an alternative scenario of prebiological accumulation and co‐evolution of homogeneous systems (RNA and DNA).     

大肠杆菌DNA和RNA中核苷的高效液相色谱分析

建立了用反相高效液相色谱法（RP-HPLC）测定大肠杆菌（E.coli）脱氧核糖核酸（DNA）和核糖核酸（RNA）水解产物中核苷的方法，色谱柱为Supelco Inc.c18反相柱，254nm和280nm紫外吸收波长同时检测，分析条件经选择在流速为1mL/min，流动相A（2.5%甲醇 0.01mol/L KH2PO4,PH4.6)28min，流动相B（8.0%甲醇 0.01mol/L KH2PO4，PH4.6)37min，柱温为20℃条件下65min可实现核苷的完全分离，回收率为80.19%-99.31%，相对标准偏差RSD为0.86%-2.62%，该方法具有高灵敏度、高选择性的特点，得到的结果与十分满意。