排序方式: 共有36条查询结果,搜索用时 15 毫秒
1.
Yang XiaoHai Wang Lei Wang KeMin Tan WeiHong Tang HongXing Meng XiangXian Guo QiuPing 《科学通报(英文版)》2008,53(2):204-208
In recent years, specific detection of proteins is one of the hot issues about aptamers in proteomics. Here we reported a simple, sensitive and specific proximity-dependent protein assay with dual DNA aptamers. Thrombin was used as the model protein, and two aptamer probes with complementary sequence at 3′-end were designed for the two distinct epitopes of the protein. Association of the two aptamers with thrombin resulted in stable hybrids due to the proximity of 3′-end, then polymerase reaction was induced. The amount of obtained dsDNA was indicated using the fluorescence dye Sybr Green I. The results showed that the initial velocity of polymerase reaction had a positive correlation with concentration of thrombin. The advantages of this dual-aptamer-based approach included simple and flexible design of aptamer probes, high selectivity and high sensitivity. The detection limit was 6.9 pmol/L. 相似文献
2.
Peptide aptamers have emerged as powerful new tools for molecular medicine. They can specifically bind to and functionally
inactivate a given target molecule under intracellular conditions. Typically, peptide aptamers are generated by screening
a randomized peptide expression library, displayed from the Escherichia coli thioredoxin A (TrxA) protein. Here, we transferred peptide moieties from defined TrxA-based peptide aptamers to alternative
scaffold proteins, such as the green fluorescent protein and staphylococcal nuclease. Yeast and mammalian two-hybrid assays
as well as in vitro binding analyses show that the TrxA scaffold can be a major determinant for the binding of peptide aptamers.
In addition, we demonstrate that TrxA can correctly display peptide sequences that correspond to the binding domains of natural
interaction partners. Therefore, sequence analyses of TrxA-based peptide aptamers, isolated by two-hybrid screening from randomized
expression libraries, should also be useful to find cellular binding partners for a given target protein, by homology.
Received 1 August 2002; received after revision 17 September 2002; accepted 19 September 2002
RID="*"
ID="*"Corresponding author. 相似文献
3.
MUC1粘蛋白是一种高糖化、高分子量的糖蛋白,其在乳腺癌细胞中高度异常表达,其特异性高于组织多肽抗原,敏感性高于癌胚抗原,因此MUC1在乳腺癌诊断中具有很高的临床应用价值,而建立高灵敏的MUC1蛋白定量检测方法对临床诊断具有重要的意义.该研究建立了基于核酸适配体-滚环扩增(RCA)和氧化石墨烯-荧光共振能量转移(GO-FRET)技术的MUC1黏蛋白定量检测技术,实现了MUC1粘蛋白准确、灵敏的定量检测.结果表明,该方法定量检测线性范围为50~1 000 pg/mL,检测限为28.05 pg/mL,定量限为45.57 pg/mL,在人血样品中的回收率为96%~104%. 相似文献
4.
核酸适配体(Aptamer)是指利用指数富集配体系统进化(Systematic evolution of ligands by exponential enrichment,SELEX)技术筛选得到的能特异性识别和结合靶标的ssDNA或RNA分子,具有高亲和力和高特异性的特点,在快速检测和靶向治疗方面应用广阔。近年来,水生生物细菌和病毒性病原疾病的频繁暴发,严重制约了水生态的健康以及水产养殖业的迅速发展。核酸适配体作为一种新型的识别分子,在水生生物病原的应用上也已取得了一些进展。本文对水生生物细菌和病毒性病原核酸适配体的筛选,及其在检测和治疗领域的研究现状进行综述,并对该领域核酸适配体技术的应用方向进行展望。 相似文献
5.
小檗碱是一种价格低廉且生物安全性良好的小分子化合物,筛选能与小檗碱互相作用的核酸片段,旨在开发新型遗传元件并拓展其在生物技术领域的应用。利用指数富集的配基系统进化技术(SELEX),经8轮实验从随机单链DNA文库中筛选出4条能与小檗碱高度特异性结合的核酸适配体。对其中两条重现性较好的适配体进行结构模拟分析表明,适配体序列富含鸟嘌呤和胞嘧啶且多形成茎环及G-四链体结构。亲和力测定结果显示,其中的2条核酸适配体与小檗碱作用的解离常数Kd都处于nmol级,表现出较高的亲和力,适配体S1的Kd值最小,仅为523 nmol/L,与小檗碱亲和力最大。 相似文献
6.
以三聚氰胺作为检测目标,设计合成了能与三聚氰胺特异性结合的适配体,选取表面活性剂邻苯二甲酸二乙二醇二丙烯酸酯(PDDA),将纳米金比色法与适配体的分子识别相结合,确立了一种简单、高效、快速检测三聚氰胺的新方法.在三甲基硅丙磺酸(pH 7.0)缓冲溶液条件下,此反应体系的最佳条件为25nmol/L适配体,0.44nmol/L PDDA,反应时间为20min,通过可见光吸收光谱可检测的检测下限为34nmol/L.在牛奶样品的检测中,三聚氰胺的加标回收率为90%~120%,可以满足牛奶样品中三聚氰胺残留的精确检测要求. 相似文献
7.
建立了一个集成DNA聚合和RNA转录过程,能够重复产生RNA适体片段,并结合孔雀石绿产生荧光信号的分子机器,并探索了此分子机器在检测DNA方面的应用.转录产生的大量孔雀石绿适体序列被释放到溶液中,并可以与孔雀石绿结合产生荧光,实现信号的放大.85μL体系中的聚合转录的最适条件为:DNA聚合酶10 IU(0.118 IU·μL-1),RNA聚合酶60 IU(0.706 IU·μL-1),反应时间为3h.在上述条件下,荧光信号随着引发DNA用量的增加而增大. 相似文献
8.
张红鸽;王敏娟;漆红兰;高强;张成孝 《西北大学学报(自然科学版)》2011,41(4):617-622
目的研究一种简单、高灵敏、高选择性检测溶菌酶的比色方法。方法根据比色法进行定性分析,利用紫外可见分光光度法,根据610nm和525nm波长下吸光度之比(A610/A525)进行溶菌酶的定量检测。结果 A610/A525与溶菌酶浓度在10~100 nmol/L之间呈良好的线性关系,线性方程为A610/A525=0.007 3C(nmol/L)+0.254(r2=0.994 5),检出限为3.4 nmol/L(S/N=3)。结论所建立的检测溶菌酶的比色方法简单、选择性好、灵敏度高,可作为一种适体传感器的模型。 相似文献
9.
A magnetically assisted fluorescence ratiometric technique has been developed for adenosine deaminase assays with high sensitivity using water-soluble cationic conjugated polymers (CCPs). The assay contains three elements: a biotin-labeled aptamer of adenosine (biotin-aptamer), a signaling probe single-stranded DNA-tagged fluorescein at terminus (ssDNA-Fl) and a CCP. The specific binding of adenosine to biotin-aptamer makes biotin-aptamer and ssDNA-Fl unhybridized, and the ssDNA-Fl is washed out after streptavidin-coated magnetic beads are added and separated from the assay solution under magnetic field. In this case, after the addition of CCP to the magnetic beads solution, the fluorescence resonance energy transfer (FRET) from CCP to fluorescein is inefficient. Upon adding adenosine deaminase, the adenosine is converted into inosine, and the biotin-aptamer is hybridized with ssDNA-Fl to form doubled stranded DNA (biotin-dsDNA-Fl). The ssDNA-Fl is attached to the magnetic beads at the separation step, and the addition of CCP to the magnetic beads solution leads to efficient FRET from CCP to fluorescein. Thus the adenosine deaminase activity can be monitored by fluorescence spectra in view of the intensity decrease of CCP emission or the increase of fluorescein emission in aqueous solutions. The assay integrates surface-functionalized magnetic particles with significant amplification of detection signal of water-soluble cationic conjugated polymers. Supported by the “100 Talents” Program of the Chinese Academy of Sciences, and the National Natural Science Foundation of China (Grant No. 20574073) 相似文献
10.
核酸适体是由指数富集配体系统进化技术(SELEX)筛选获得的能够与靶标物质特异性、高亲和力结合的单链DNA或RNA.能够被核酸适体识别的靶标物质包括各种小分子物质、大分子蛋白质、细胞等,广泛应用于生物医学研究各领域.为此,对核酸适体的筛选制备技术、对生物医学靶标物质的检测方法以及在疾病的靶向诊断和治疗中的研究进展进行了综述,并对其在生物医学领域中的应用进行了展望. 相似文献