目的使用R6G-ddATP作为双脱氧荧光底物建立单碱基末端延伸(SNaPShot)-凝胶荧光法快速检测3种高危型人乳头瘤病毒(high risk human papillomavirus,HR-HPV)(HPV18、HPV33、HPV35)基因型。方法使用HPV质控品作为样本,R6G-ddATP双脱氧荧光...目的使用R6G-ddATP作为双脱氧荧光底物建立单碱基末端延伸(SNaPShot)-凝胶荧光法快速检测3种高危型人乳头瘤病毒(high risk human papillomavirus,HR-HPV)(HPV18、HPV33、HPV35)基因型。方法使用HPV质控品作为样本,R6G-ddATP双脱氧荧光试剂作为底物,首先利用通用引物对HPV进行扩增,得到第一轮扩增产物,经纯化后作为后续SNaPShot反应的模板;然后利用特异性的一步延伸引物进行SNaPShot反应,生成带有R6G荧光标记的DNA延伸产物;产物经过琼脂糖凝胶电泳,在凝胶成像仪下观察电泳结果,通过不同的一步延伸引物对HPV进行分型。每个样本均重复检测3次,并与DNA测序结果进行比较。结果优选的SNaPShot反应的退火温度为55℃;仅需3 h即可对HPV进行分型;在该最适条件下使用R6G-ddATP/SNaPShot-凝胶荧光法检测3种HPV基因型,检测结果与测序结果一致。结论成功建立了3种HR-HPV基因型的微量检测方法——R6G-ddATP/SNaPShot-凝胶荧光法,可用于HPV基因型的快速检测。展开更多
One of the rapidly growing fields of nanotechnology is its manipulation of laser dyes’properties using nanoparticles and nanostructures due to its various applications,ranging from biomedical imaging to green energy....One of the rapidly growing fields of nanotechnology is its manipulation of laser dyes’properties using nanoparticles and nanostructures due to its various applications,ranging from biomedical imaging to green energy.Silver nanoparticles(Ag NPs)of various concentrations and nanostructures with silver nanowire(Ag NW)were prepared using an electrical exploding wire technique(EEW)and was mixed with a fixed concentration of R6G dye.The behavior of energy transfer from the dye molecules(R6G)to nanomaterials(Ag NPs or plasmonic nanostructures)was examined using fluorescence spectra.The experimental results showed that the fluorescence intensity quenched with increasing concentration and density number of Ag NPs.The distance between the dye molecules and the nanostructures was studied,which was found to decrease as the concentration and density number of Ag NPs increased in the mixture.The energy transfer efficiency of nanostructures was compared.It was obtained that nanostructure(Ag NW@PDA@Ag NPs)achieved the best energy transfer efficiency of 85%.Our results indicated that this nanostructure could sense a distance around the metal nanoparticles(≈27.2 nm);thus the nanoparticle-based surface energy transfer(NSET)mechanism is dominated rather than Förster resonance energy transfer(FRET)mechanism.This process is affected by concentration increasing of Ag NPs and coated morphology of Ag NWs by polydopamine(PDA)layer decorated by Ag NPs.The findings can be utilized in the large field of bio diagnostics and biochemistry.Regardless of bio-applications,the quenching mechanisms and rates are also of interest for SERS,(dye-sensitized)solar cells or nanooptics.However,we see the best potential in bio-sensing by managing the quenching rate by adjusting the shape or the concentration of nanostructures.展开更多
文摘目的使用R6G-ddATP作为双脱氧荧光底物建立单碱基末端延伸(SNaPShot)-凝胶荧光法快速检测3种高危型人乳头瘤病毒(high risk human papillomavirus,HR-HPV)(HPV18、HPV33、HPV35)基因型。方法使用HPV质控品作为样本,R6G-ddATP双脱氧荧光试剂作为底物,首先利用通用引物对HPV进行扩增,得到第一轮扩增产物,经纯化后作为后续SNaPShot反应的模板;然后利用特异性的一步延伸引物进行SNaPShot反应,生成带有R6G荧光标记的DNA延伸产物;产物经过琼脂糖凝胶电泳,在凝胶成像仪下观察电泳结果,通过不同的一步延伸引物对HPV进行分型。每个样本均重复检测3次,并与DNA测序结果进行比较。结果优选的SNaPShot反应的退火温度为55℃;仅需3 h即可对HPV进行分型;在该最适条件下使用R6G-ddATP/SNaPShot-凝胶荧光法检测3种HPV基因型,检测结果与测序结果一致。结论成功建立了3种HR-HPV基因型的微量检测方法——R6G-ddATP/SNaPShot-凝胶荧光法,可用于HPV基因型的快速检测。
文摘One of the rapidly growing fields of nanotechnology is its manipulation of laser dyes’properties using nanoparticles and nanostructures due to its various applications,ranging from biomedical imaging to green energy.Silver nanoparticles(Ag NPs)of various concentrations and nanostructures with silver nanowire(Ag NW)were prepared using an electrical exploding wire technique(EEW)and was mixed with a fixed concentration of R6G dye.The behavior of energy transfer from the dye molecules(R6G)to nanomaterials(Ag NPs or plasmonic nanostructures)was examined using fluorescence spectra.The experimental results showed that the fluorescence intensity quenched with increasing concentration and density number of Ag NPs.The distance between the dye molecules and the nanostructures was studied,which was found to decrease as the concentration and density number of Ag NPs increased in the mixture.The energy transfer efficiency of nanostructures was compared.It was obtained that nanostructure(Ag NW@PDA@Ag NPs)achieved the best energy transfer efficiency of 85%.Our results indicated that this nanostructure could sense a distance around the metal nanoparticles(≈27.2 nm);thus the nanoparticle-based surface energy transfer(NSET)mechanism is dominated rather than Förster resonance energy transfer(FRET)mechanism.This process is affected by concentration increasing of Ag NPs and coated morphology of Ag NWs by polydopamine(PDA)layer decorated by Ag NPs.The findings can be utilized in the large field of bio diagnostics and biochemistry.Regardless of bio-applications,the quenching mechanisms and rates are also of interest for SERS,(dye-sensitized)solar cells or nanooptics.However,we see the best potential in bio-sensing by managing the quenching rate by adjusting the shape or the concentration of nanostructures.
