硅烷法固定ssDNA制作的压电式DNA传感器

利用硅烷法将ssDNA 固化T方向切割、3. 58MHz 和10MHz 石英晶体金电极上,构建了压电DNA传感器,建立了快速检测葡萄球菌肠毒素B(SEB) 基因的方法。优化了固定SEB 基因的条件,并用动态法、静态法和斑点杂交法进行了固化效果观察。结果表明:该传感器2 h 后,有较好响应;对不同长度DNA 探针固化效果进行比较,169 bp (即169 个碱基) SEB 探针固定和杂交效果较好;利用0. 5mol/ LNaOH进行电极洗脱,时间为45min ,电极可以再生,且传感器可以重复使用3 次;利用1. 2mol/ L NaOH和1. 2mol/ L HCl洗涤电极,该电极可多次再利用;固化好的电极于4 ℃条件下可以保存三周之久。
关键词: 压电式DNA 传感器; 葡萄球菌肠毒素B ; 3 - 氨基丙基三乙氧基甲硅烷
Abstract : A rapid method has been developed to monitor Staphylococcal enterotoxin B(SEB) gene by using piezoeletric DNA sensor system. AT2cut crystals ,with a based resonant frequency of 3. 58MHz and 10MHz ,were modified by 32 aminopropyltriethoxysilane(32APTES) treatment and then single stranded SEB gene probe was electro2blotted onto the
surface of crystal electrodes. The optimization condition of immobilization for piezoeletric DNA sensor was researched. Im2 mobililzed efficiency was observed by active methods , passive methods and dot hybridization method. There is better re2 sponse of the piezoelectric DNA sensor after DNA hybridization for 2 h. Hybridization effect of 169 bp SEB gene probe was better comparing with the other different length of SEB gene probe. The sensor could be repeatedly used ,the complemen2 tary ssDNA of the SEB DNA probe could be removed from the crystal by washing with 0. 5mol/ L NaOH denaturant. After the hybridized electrode was immersed in 1. 2mol/ L NaOH and then 1. 2 mol/ L HCl ,all substances could be removed
from the crystal surface ,and the quartz crystal could be reused for many times. The 32APTES2coated ssDNA electrode was stable for over three weeks when stored at 4 ℃.
Key words : piezoelectric DNA sensor ; staphylococcal enterotoxin B (SEB) ; 32aminopropyltriethoxysilane (32APTES)