黄连素标记的脱氧核糖核酸荧光毛细生物传感器的研制及应用
Preparation and Application of DNA Fluorescence Capillary Biosensor Labelled with Berberine
摘 要
以黄连素作为荧光标记物,对脱氧核糖核酸荧光毛细生物传感器进行了研究.基于荧光毛细分析法,以毛细管作为脱氧核糖核酸探针的固定载体,制成脱氧核糖核酸荧光毛细生物传感器,荧光毛细生物传感器吸入互补靶脱氧核糖核酸并进行杂交,通过黄连素染色后,检测杂交产物的荧光强度,实现对靶脱氧核糖核酸的定性和定量分析.研究结果为样品用量12 μL,靶脱氧核糖核酸浓度在2~10 μmol·L-1范围内与荧光强度呈线性关系,线性回归方程为y=9.52 x+9.22,相关系数为0.991 2,检出限为1.16 μmol·L-1.用荧光毛细生物传感器测定靶脱氧核糖核酸操作简便,试样、试剂用量少;测定成本极低,能大大减少环境污染.
DNA荧光毛细生物传感器
荧光标记物
黄连素
DNA
DNA fluorescence capillary biosensor
Fluorescence labelling substance
Berberine
Abstract
In preparing DNA fluorescence capillary biosensor(DNA-FCB),capillary was used as carrier to immobilize DNA and complementary target DNA was sucked into the capillary for the hybridization to form spiral dual chain DNA in which berberine was embedded as the fluorescence labelling substance.By measuring the fluorescence intensity with a fluorospectrophotometer,the target DNA was detected qualitatively and quantitatively.Optimum conditions for the determination were found as follows: ① volume of sample solution taken: 12 μL;② range of linearity and linear regression equation: 2-10 μmol·L-1 of target DNA,y=9.52 x+9.22(r=0.991 2);③ detection limit: 1.16 μmol·L-1 of target DNA.The method of determination of target DNA with this biosensor was proved to be simple and convenient in operation,low in consumption of reagents and in cost for determination,as well as profitable to environmental protection.
中图分类号 O657.31
所属栏目 试验与研究
基金项目 四川大学振兴计划科研启动基金资助项目(0082204127092)
收稿日期 2007/12/24
修改稿日期
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备注杨全玉(1977-),女,满族,辽宁人,硕士研究生,研究方向为DNA生物传感器.
引用该论文: YANG Quan-yu,LI Yong-sheng,GAO Xiu-feng,WANG Yan-jun,JIN Ke,ZHAO Rui. Preparation and Application of DNA Fluorescence Capillary Biosensor Labelled with Berberine[J]. Physical Testing and Chemical Analysis part B:Chemical Analysis, 2008, 44(6): 491～494
杨全玉,李永生,高秀峰,王艳君,金科,赵瑞. 黄连素标记的脱氧核糖核酸荧光毛细生物传感器的研制及应用[J]. 理化检验－化学分册, 2008, 44(6): 491～494
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参考文献
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【3】Natalia C T,Xie H,Xie F,et al.Direct detection of DNA with an electrocatalytic threading intercalator[J].Anal Chem,2005,77(1):126-134．
【4】Wang J,Palecek E,Nielsen P E,et al.Peptide nucleic acid probes for sequence-specific DNA biosensor[J].J Am Chem Soc,1996,118:7667-7670．
【5】Tombelli S,Sacco C,Turner A P F,et al.A DNA piezoelectric biosensor assay coupled with a polymerase chain reaction for bacterial toxicity determination in environmental samples[J].Anal Chim Acta,2000,418(1):1-9．
【6】Piunno P A E,Krull U J,Hudson R H E,et al.Fiber optic biosensor for fluorimetric detection of DNA hybridization[J].Anal Chim Acta,1994,288(3):205-214．
【7】姜广奋,陈润生,阎宏,等.用于基因探测的光纤DNA传感器及其阵列的一种新的制备方法[J].中国科学:C辑,2000,30(5):461-466．
【8】翟俊辉,黄惠杰.光纤生物传感器用于核酸的特异性检测[J].分析化学,2003,31(1):34．
【9】杜江燕,黄晓华,徐飞,等.硫堇与DNA分子作用机理的光谱研究[J].光谱学与光谱分析,2005,25(9):1435．
【10】李永生,高秀峰.荧光毛细反应装置及荧光毛细分析法:中国,200510021542,8[P],2005-10-2．
【11】高秀峰,李永生,姜艳霞.Cy-5标记脱氧核糖核酸荧光毛细分析法的研究[J].分析化学,2006,34(9):S220-S222．
【12】李永生,高秀峰.一种用于化学分析的新方法-荧光毛细分析法[J].光谱学与光谱分析,2007,27(8):1565-1569．
【13】Li Y S,Gao X F.Determination of various alcohols based on a new immobilized enzyme fluorescence capillary analysis[J].Anal Chim Acta,2007,588(1):140-146．
【14】Lunn G,Sansone E B.Ethidium bromide: destruction and decontamination of solutions[J].Anal Biochem,1987,162(2):453．
【15】海志杰,沈宏伟,邓军卫.两种核酸染料的应用比较研究[J].实用预防医学,2004,11(5):941．
【16】Zhang X B,Li Z Z,Guo C C,et al.Porphyrin-metalloporphyrin composite based optical fiber sensor for the determination of berberine[J].Anal Chim Acta,2001,439(1):65-71．
【17】Gong G Q,Zong Z X,Song Y M.Spectrofluorometric determination of DNA and RNA with berberine[J].Spectrochim Acta Part A,1999,55(9):1903-1907．
【18】赵长春,于俊生,章杰兵.药物与DNA的相互作用研究Ⅰ:小檗碱作为DNA的发光开关及pH的影响[J].无机化学学报,1997,13(3):325-329．
【19】何群,赵雨杰.一种新的DNA芯片的封闭方法[J].遗传,2004,26(3):361-363．
【20】吴成铁,陈敏,李长敏,等.用多聚-L-赖氨酸活化生物芯片玻璃片基的研究[J].材料导报,2003,17(4):81-83．
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