生物素-亲和素-DNA纳米颗粒信号放大检测载体的构建

目的构建用于检测细菌、病毒等病原体DNA的生物素-亲和素纳米颗粒信号放大载体。方法设计并合成两端和一端标记生物素的寡核苷酸探针(2B/1B-DNA),与抗生蛋白链菌素葡聚糖(Poly-STV)通过生物素与亲和素作用,偶联形成大分子纳米网状颗粒,并筛选最佳探针类型及其长度、2B-DNA和Poly-STV的浓度及比例和反应条件。结果2B-DNA和Poly-STV形成纳米颗粒信号放大载体的最佳浓度分别为1μmol/L和5μmol/L,最佳浓度比为1∶5,2B-DNA长度为60bp,缓冲液为Buffer B,反应条件为55℃,800r/min,20min。结论已成功构建了生物素-亲和素大分子纳米颗粒,可作为DNA检测信号放大技术的备选载体。

Construction of Biotin-Streptavidin-DNA Nanoparticle Signal Amplification Carrier for Nucleic Acid Assay

Objective To construct a biotin-streptavidin-DNA nanoparticle signal amplification carrier for assay of DNA in pathogens such as virus and bacterium.Methods Mono-and bis-biotinylated oligonucleotide probes(1B /2B-DNA)were designed and synthesized,then coupled to streptavidin-labeled dextran(Poly-STV)through the interaction of biotin and streptavidin to construct a macromolecular nanoparticle signal amplification carrier.The kind and length of probe,the concentrations of 2B-DNA and Poly-STV,the ratio of 2B-DNA to Poly-STV as well as reaction condition were optimized.Results The optimal concentrations of 2B-DNA and Poly-STV for construction of the carrier were 1 and 5 μmol /L respectively,the optimal ratio of 2B-DNA to Poly-STV was 1:5,the optimal length of 2B-DNA was 60 bp,the optimal reaction condition was 55℃,800 r /min for 20 min.Conclusion The biotin-streptavidin-macromolecular nanoparticles were successfully constructed,which might be used as a candidate carrier for DNA signal amplification detection technology.