现场光化学荧光探针法测定DNA──以9，10－蒽醌－2－磺酸钠为光化学荧光探针

现场光化学荧光探针法测定ＤＮＡ──以９，１０－蒽醌－２－磺酸钠为光化学荧光探针李文友，许金钩，郭祥群，朱庆枝，赵一兵（材料和生命过程分析科学开放研究实验室，厦门大学化学系，厦门，３６１００５）关键词ＤＮＡ，光化学荧光探针，９，１０－蒽醌－２－磺酸钠光...     

吖啶橙(AO)-藏红T(ST)荧光探针测定DNA的研究

简述了以荧光能量转移本系测定ＤＮＡ方法的发展过程，同时对吖啶（ＡＯ）－藏红Ｔ（ＳＴ）荧光探针测定ＤＮＡ的条件进行分析，证明了该法灵敏度高、线性范围宽、检出限低、选择性好、抗干扰能力强，该法为微曦ＤＮＡ测定分析工作提供了一个新的途径。     

阿达玛变换显微图像分析系统在乳腺肿瘤细胞DNA倍性分析…

以正常人外周静脉血淋巴细胞为标准二倍体细胞，以吖橙为细胞ＤＮＡ荧光探针，用阿达玛变换显微图象分析仪测定了冰例乳腺肿瘤的细胞ＤＮＡ倍性与面积呈显著正相关。     

荧光能量转移体系在生化分析中的应用-吖啶橙(AO)-中性红(NR)荧光探针测定DNA研究

本文简述了以荧光能量转移体系测定ＤＮＡ方法的发展过程,对近几年这一领域的新成果作了介绍;同时对吖啶橙（ＡＯ）－中性红ＮＲ为荧光探针测定ＤＮＡ的条件进行分析,证明了该法具有灵敏度高、线性范围宽、检出限低、选择性好、抗干扰能力强等,该法为微量ＤＮＡ定量分析提供了一个新的途径。     

铽（Ⅲ）探针法研究一些金属离子与DNA的作用

本文首次选用Ｔｂ（Ⅲ）作为荧光探针，研究了ｃｄ（Ⅱ）、Ｚｎ（Ⅱ）、Ｃａ（Ⅱ）、Ｍｇ（Ⅱ）与ＤＮＡ反应的结合部位。结果表明：Ｃｄ（Ⅱ）、Ｚｎ（Ⅱ）在ＤＮＡ上有两种结合部位，即ＤＮＡ双螺旋外侧的磷酸基和内侧的碱基鸟漂呤上的Ｎ－Ｔ。而ｃａ（Ⅱ）、Ｍｇ（Ⅱ）在ＤＮＡ上只有一个结合部位即磷酸基。说明Ｔｂ（Ⅲ）不仅可以作为ＤＮＡ构象及ＤＮＡ单链成分、不配对碱基残余的探针，而且可以用它探测一些金属离子与ＤＮＡ反应的结合部位。     

核苷酸,聚核苷酸和核酸猝灭Tb^3＋—钛铁试剂络合物荧光的…

研究了核苷酸，聚核苷酸和核酸对Ｔｂ＾３＋－钛铁试剂（ＴＲ）络合物的荧光猝灭机理，认为荧光猝灭过程是核苷酸，聚核苷酸和核酸分子中的磷酸基组分为与ＴＲ竞争Ｔｂ＾３＋离子，生成实验条件下无荧光的二元络合物的静态猝灭过程；用Ｔｂ＾３＋－ＴＲ络合物荧光探针研究了ＤＮＡ嵌入剂和金属离子与ＤＮＡ相互作用的实验结果说明这一机理是合理的。     

荧光法研究手性金属配合物与DNA的作用机理

以溴化乙锭为荧光探针，研究手性金属配合物［Ｎｉ（ｐｈｅｎ）３］＾＋和［Ｆｅ（ｐｈｅｎ）３］＾２＋与ＤＮＡ的反应机理。结果表明，配合物与ＤＮＡ作用存在插入和静电结合两种模式，即部分菲咯啉配体插入ＤＮＡ双螺旋碱基对中，同时带正电荷的配合物与ＤＮＡ的磷酸基团发生静电结合。     

DNA探针在羧基聚苯乙烯微珠表面的微阵列研究

报道在１－乙基－３－３－（３－二甲基氨丙基）－碳化二亚胺（ＥＤＣ）偶链剂存在下,５’－ＮＨ_２单末端和５’－Ｔｅｘ／３’－ＮＨ_２双末端修饰ＤＮＡ探针在１０μｍ 羧基功能微珠表面的微阵列特性．研究表明,ＤＮＡ探针在微珠表面的微阵列能力决定于溶液介质的ｐＨ,ＤＮＡ探针浓度和微珠总表面积．优化条件下, ２０ ｍｅｒ单链ＤＮＡ探针在微珠表面微阵列的最小分子间距约为１４ ｎｍ,而２０ ｍｅｒ双链ＤＮＡ分子微阵列的最小分子间距约为２７ ｎｍ．如果ＤＮＡ探针序列长度由 ２０增加到 ３５ ｍｅｒ,探针在微珠表面的微阵列密度降低．机理研究表明,ＤＮＡ探针在羧基功能微珠表面的微阵列方式是探针分子近平行于微珠表面．     

中性红作为DNA作用方式光谱探针的研究

以溴乙锭（ＥＢ）为探针在研究吩噻嗪药物与ＤＮＡ间相互作用的基础上,用中性红（Ｎｅｕｔｒａｌ Ｒｅｄ,ＮＲ）代替ＥＢ为ＤＮＡ作用方式对光谱探针的可行性进行了研究。结果表明,ＮＲ作为ＤＮＡ作用方式光谱探针与ＥＢ具有可比性,且由于其低毒性和高稳定性而具有较好的应用潜力。     

人的基因组及DNA序列分析：过去,现在及将来（下）

对ＤＮＡ序列分析方法如毛细管电泳、阵列毛细管电泳、超薄层板电泳作了详细评述。并对正在研究开发不用电泳分离的直接测序新技术和新方法，如质谱法、原子探针法（扫描隧道显微镜、原子力显微镜）、杂交法、流动式单分子荧光检测法等进行了评论。     

Twin probes as a novel tool for the detection of single-nucleotide polymorphisms

Single-nucleotide polymorphisms (SNPs) are the most common form of DNA sequence variation. There is a strong interest from both academy and industry to develop rapid, sensitive and cost effective methods for SNP detection. Here we report a novel structural concept for DNA detection based on fluorescence dequenching upon hybridization. The so-called "twin probe" consists of a central fluorene derivative as fluorophore to which two identical oligonucleotides are covalently attached. This probe architecture is applied in homogeneous hybridization assays with subsequent fluorescence spectroscopic analysis. The bioorganic hybrid structure is well suited for sequence specific DNA detection and even SNPs are identified with high efficiency. Additionally, the photophysical properties of the twin probe were investigated. The covalent attachment of two single stranded oligonucleotides leads to strong quenching of the central fluorescence dye induced by the nucleobases. The twin probe is characterized by supramolecular aggregate formation accompanied by red-shifted emission and broad fluorescence spectra.     

用于细胞核成像的2,7-BHVC双光子荧光探针

分别合成了2,7-双(1-羟乙基-4-乙烯基吡啶) 咔唑碘盐(2,7-BHVC)与3,6-BHVC, 进行了荧光性能和细胞核成像测试, 结果表明, 2,7位的Ф×δ是3,6位的2.8~3.6倍, 染色实验也证实了2,7-BHVC有特定成像细胞核的能力.     

一种新型发光探针与DNA相互作用的研究

本文以稳态荧光光谱、紫外-可见吸收光谱、荧光偏振、热变性、阴离子猝灭等手段,研究了一种具有强电荷转移能力的化合物2,7-二[(N-乙基咔唑-3-基)丙烯-1-酮基]芴与DNA的相互作用。研究结果表明,2,7-二[(N-乙基咔唑-3-基)丙烯-1-酮基)芴与DNA的作用方式是混合模式,以嵌插作用为主,同时存在沟槽相互作用,其咔唑基团可能插入到DNA的碱基对之间,结合常数K为8 123.48mol/L,结合位点n为0.71。该发光探针灵敏度高,结合稳定。     

Monitoring DNA structures by dual fluorescence of pyrene derivatives

We have developed a nucleotide modified by a pyrene derivative with dual fluorescence. The dual fluorescence of the fluorophore, which was incorporated into DNA, was effectively controlled at ambient temperature according to DNA structural status. Our nucleoside with dual fluorescence is effective as a conceptually new probe for monitoring DNA hybridization by the color change without multilabeling with fluorescent dyes.     

A homogeneous fluorescence sensing platform with water-soluble carbon nanoparticles for detection of microRNA and nuclease activity

Based on the high efficiency of fluorescence quenching and the different affinities of water-soluble carbon nanoparticles (CNPs) towards single-stranded DNA (ssDNA) and double-stranded DNA/RNA hybrid, a novel, rapid and cost-effective assay for detection of microRNA and nuclease activity was developed. The fluorescein-labeled ssDNA probe (FAM-P) could be adsorbed on the surface of CNPs through π-π stacking interaction giving rise to fluorescence quenching. By introduction of microRNA complementary to the DNA probe, the double-stranded DNA/miRNA hybrid could be formed and released from the surface of CNPs resulting in the fluorescence recovery. Thus, microRNA was successfully detected in homogenous fashion without any amplification or enzyme-involving reactions. Moreover, we demonstrated that the nuclease activities of RNase H and DNase I could also be sensitively monitored by using CNPs based on the fluorescence changing of the DNA probe. So, the CNPs provide an excellent homogeneous sensing platform for studying molecular diagnosis and therapeutics.     

A DNA hybridization detection based on fluorescence resonance energy transfer between dye-doped core-shell silica nanoparticles and gold nanoparticles

A novel and efficient method to evaluate the DNA hybridization based on a fluorescence resonance energy transfer (FRET) system, with fluorescein isothiocyanate (FITC)-doped fluorescent silica nanoparticles (SiNPs) as donor and gold nanoparticles (AuNPs) as acceptor, has been reported. The strategy for specific DNA sequence detecting is based on DNA hybridization event, which is detected via excitation of SiNPs-oligonucleotide conjugates and energy transfer to AuNPs-oligonucleotide conjugates. The proximity required for FRET arises when the SiNPs-oligonucleotide conjugates hybridize with partly complementary AuNPs-oligonucleotide conjugates, resulting in the fluorescence quenching of donors, SiNPs-oligonucleotide conjugates, and the formation of a weakly fluorescent complex, SiNPs-dsDNA-AuNPs. Upon the addition of the target DNA sequence to SiNPs-dsDNA-AuNPs complex, the fluorescence restores (turn-on). Based on the restored fluorescence, a homogeneous assay for the target DNA is proposed. Our results have shown that the linear range for target DNA detection is 0-35.0 nM with a detection limit (3σ) of 3.0 picomole. Compared with FITC-dsDNA-AuNPs probe system, the sensitivity of the proposed probe system for target DNA detection is increased by a factor of 3.4-fold.     

荧光法研究抗癌药物更生霉素D与小牛胸腺DNA的作用机理

以溴化乙锭(ethidiumbromide,EB)为荧光探针,研究了更生霉素D(actinomyinD,ACTD)与小牛胸腺DNA(calfthymusDNA,CTDNA,以下简称DNA)的作用机理。对荧光光谱、荧光偏振、Scatchard图、DNA热变性曲线、DNA盐效应曲线等研究表明,ACTD与DNA之间主要存在嵌入和沟槽两种作用,并证明ACTD与DNA磷酸基之间不存在静电作用。     

Determination of DNA using sodium 9,10-anthraquinone-2-sulfonate as an in situ photochemical fluorescence probe

An in situ photochemical fluorescence probe method for the determination of DNA with sodium 9,10-anthraquinone-2-sulfonate (AQS) as a photochemical fluorescence probe was developed. It was based on the conversion of AQS into an intensively fluorescent product by irradiating with UV radiation. The photochemical reaction is retarded by DNA. The determination can be carried out by measuring the fluorescence intensity at a fixed time. The calibration graph was linear in the range 0–80 ng ml⁻¹ calf thymus (CT) DNA (r = 0.9991), the limit of detection was 3.2 ng ml⁻¹ CT DNA (n = 9). The kinetic behaviour of the photochemical reaction and the effects of experimental conditions were investigated and discussed in detail. The results of absorption spectra and competitive binding experiments suggested the interaction between AQS and DNA to be intercalative.     

Application of Single—labelled Probe—primer in PCR Amplification to the Detection of Hepatitis B Virus DNA

A new method based on the incorporation of a single-lablled probe-primer into polymerase chain reaction(PCR) for the detection of PCR-amplified DNA in a closed system is reported.The probeprimerc consists of a specific probe sequence on the 5‘‘‘‘‘‘‘‘-end and a primer sequence on the 3‘‘‘‘‘‘‘‘-end.A flurophore is located at the 5‘‘‘‘‘‘‘‘end.The primeR-quencher is an oligonucleotide,which is complementary to the probe sequence of probe-primer and labelled with a quencher at the 3‘‘‘‘‘‘‘‘-end.In the duplex formed by probe-primer and primer-quencher.the fluorophore and quencher are kept in close proximity to each other.Therefore the fluorescence is quenched.During PCR amplificatio,the specific probe sequence of probeprimer binds to its complement within the same strand of DNA,and is cleaved by Taq DNA polymerase,resulting in the restoration of fluorescence.This system has the same energy transfer mechanism as molecular beacons,and a good quenching effciency can be ensured.Following optimization of PCR conditions,this method was used to detect hepatitis b virus(HBV) dna in patient sera.This technology eliminates the risk of carry-over contamination,simplifies the amplification assay and opens up new possibilities for the real-time detection of the amplified DNA.