河豚毒素DNA适配子核心区域的识别

用DNA folding form在线软件分析河豚毒素DNA适配子A3、G10的二级结构,根据二级结构,分别合成适配子A3、G10的各4个区域的5’端地高辛标记的序列。用丙烯酸-环氧乙烷珠子固定河豚毒素,测定抗地高辛碱性磷酸酶的适宜稀释倍数,用地高辛-抗地高辛碱性磷酸酶系统检测各区域与河豚毒素的亲和力,识别出适配子的核心区域。结果表明,抗地高辛碱性磷酸酶的适宜工作浓度为2×104倍稀释;合成的适配子A3的4个区域中,A3-2区域与河豚毒素的亲和力最强,即适配子A3的核心区域为A3-2区域,实际上含有47个核苷酸;合成的适配子G10的4个区域中,G10-4区域与河豚毒素的亲和力最强,即适配子G10的核心区域为G10-4区域,实际上含有56个核苷酸。

Identification of Core Region of DNA Aptamers against Tetrodotoxin

The secondary structures of DNA aptamers A3 and G10 against tetrodotoxin were analyzed using DNA folding form online software. Further, digoxin labeled 5’ end sequences of four regions in each aptamer were separately synthesized. Tetrodotoxin was fixed by acrylic acid-epoxy ethane beads. The core regions of these aptamers were identified using digoxin-anti digoxin alkaline phosphatase system to detect the affinity of these various regions with tetrodotoxin, after determining the appropriate dilution ratio of anti-digoxin alkaline phosphatase. The results showed that the suitable working concentration of anti-digoxin alkaline phosphatase was 2 × 104 times dilution. The core region of aptamer A3 was A3-2 area which had the strongest affinity with tetrodotoxin among the four regions, including 47 nucleotides, while the core region of aptamer G10 was G10-4 area, including 56 nucleotides.