计时电量法用于检测急性早幼粒细胞白血病PML/RARα融合基因的研究  

Study on the Detection of PML/RARα Fusion Gene in Acute Promyelocytic Leukemia Based on Chronocoulometric Method

作　　者：汪红梅[1] 翁少煌[1] 林丽清[1] 林新华[1] 陈元仲[1] 

机构地区：[1]福建医科大学药学院药物分析系,福建福州350004

出　　处：《分析测试学报》2011年第8期872-876,共5页Journal of Instrumental Analysis

基　　金：国家自然科学基金资助项目(20675015);国家863计划资助项目(2008AA02Z433);福建省自然科学基金资助项目(2010J06011;2010J01032);福建省教育厅资助项目(JA10155);福建省高校产学研科技重点项目资助(2010Y4003)

摘　　要：以氯化六氨合钌([Ru(NH3)6]3+)为杂交指示剂,通过Au-S键的共价结合,将DNA探针固定在电极表面与互补靶序列杂交,构建了计时电量法(CC)检测急性早幼粒细胞白血病(APL)中PML/RARα融合基因的电化学DNA传感器,并通过对比杂交前后电量的变化检测互补序列及浓度。由于杂交前后与DNA链静电结合的[Ru(NH3)6]3+量不同而形成电量差值(ΔQ),随着靶序列浓度的增加,[Ru(NH3)6]3+引起的电量差值增大。在最佳实验条件下,此传感器显示了良好的特异性,能区分完全互补和错配序列,并可对融合基因进行定量。互补靶DNA序列在1.0×10-12～1.0×10-8 mol.L-1浓度范围内,ΔQ与DNA浓度的对数值呈良好的线性关系,检出限为4.0×10-13 mol.L-1。实验结果表明,以[Ru(NH3)6]3+为电化学指示剂的DNA传感器,能很好地对含PML/RARα融合基因的靶序列进行定性定量检测。An electrochemical DNA biosensor based on hexaammineruthenium（Ⅲ） chloride（[Ru（NH3）6]3＋） as a hybrid indicator was fabricated for the detection of PML/RARα fusion gene in acute promyelocytic leukemia（APL） by chronocoulometry（CC）.The DNA probes were immobilized onto the surface of electrode via Au-S covalent bond,followed by the hybridization with target DNA complementary sequences.CC method was employed to monitor the hybridization events and contrast the charge variations before and after hybridization to quantitate the target DNA complementary sequences.Owing to the different amounts of [Ru（NH3）6]3＋ caused by electrostatic combination with DNA sequences before and after hybridization,the charge difference was formed.The increase of the different charge of [Ru（NH3）6]3＋ was observed upon the hybridization of the probe with target DNA,and the charge difference of [Ru（NH3）6]3＋ was enhanced with the increase of target DNA sequence concentration.Under the optimal conditions,experimental results indicated that the biosensor showed a good specificity to distinguish the complementary sequence from different mismatch sequences and could be used to detect PML/RARα fusion gene quantitatively.The relationship between the charge difference and logarithmic（lgc） of target sequence concentrations was linear in the concentration range of 1.0×10-12-1.0×10-8 mol·L-1,and the detection limit was 4.0×10-13 mol·L-1.Therefore,as an electrochemical DNA indicator of the DNA sensor,the [Ru（NH3）6]3＋can be used to detect the complementary sequence of PML/ RARα fusion gene qualitatively and quantitatively.

关 键 词：DNA电化学杂交传感器 计时电量法 六氨合钌 PML/RARΑ融合基因 

分 类 号：O657.1[理学—分析化学] O629.74[理学—化学]