Development of a chemiluminescent reagent that recognizes guanine in nucleic acid, and its use for chemiluminescence detection of DNA

识别核酸中鸟嘌呤的化学发光试剂的研制及其在DNA化学发光检测中的应用

• 批准号: 08457640
• 负责人: KAI Masaaki
• 金额: $ 3.78万
• 依托单位: KYUSHU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08457640/
• 关键词: Nucleic acid; DNA; Chemiluminescence; Imaging detection; Arylglyoxalreagents; CCD camera; Selective detection; Hybridization; 高感度検出

A novel reagent, 3', 4', 5'-trimethoxyphenylglyoxal has been found to specifically convert guanine bases of a nucleic acid in a neutral or alkaline phosphate solution to the corresponding chemiluminescent derivatives. The chemiluminogenic derivatization technique offers a facile method of the sensitive imaging detection for a DNA probe tailing with deoxyriboguanylic acid oligomers. A cooled charge-coupled device camera is usable to detect visually the chemiluminescence emitted from the derivatized probe that was hybridized with a target DNA fragment on a Nylon membrane. The proposed method permits the quantitative detection of the probe covalently attached with a 5'- (G) _<15>TT (G) _<15>TT deoxyoligomer at 3' terminus, indicating a detection limit of 0.1 pmol on the membrane. The detection sensitivity can be increased with larger contents of deoxyriboguanylic acid in the probe molecule.

一种新的试剂，3',4',5'-三甲氧基苯乙二醛，可以特异性地将核酸中的鸟嘌呤碱基在中性或碱性磷酸盐溶液中转化为相应的化学发光衍生物。化学致发光衍生化技术为脱氧核鸟苷酸低聚物DNA探针尾端的灵敏成像检测提供了一种简便的方法。冷却电荷耦合器件摄像机可用于视觉检测衍生化探针发出的化学发光，该探针与尼龙膜上的目标DNA片段杂交。所提出的方法可以定量检测探针在3‘端共价连接5’- (G) _<15>TT (G) _<15>TT脱氧寡聚物，表明在膜上的检测限为0.1 pmol。探针分子中脱氧核鸟苷酸的含量越大，检测灵敏度越高。

项目成果

甲斐 雅亮: "グアニン塩基に特異的なルミネッセンス試薬の活用法" 蛋白質核酸酵素. 42(2). 777-783 (1997)

Masaaki Kai：“鸟嘌呤碱基特异性发光试剂的利用”蛋白质核酸酶 42(2) 777-783 (1997)。

Masaaki Kai: "Practical use of luminogenic reagents specific for guanine-containing compounds" Protein, Nucleic acid and Enzyme (Japanese). Vol42, No5. 777-783 (1997)

Masaaki Kai：“含鸟嘌呤化合物的发光试剂的实际应用”蛋白质、核酸和酶（日语）。

甲斐雅亮: "グアニン塩基に特異的なルミネッセンス試薬の活用法" 蛋白質 核酸 酵素. 42(2). 777-783 (1997)

Masaaki Kai：“鸟嘌呤碱基特异性发光试剂的利用”蛋白质核酸酶 42(2) 777-783 (1997)。

甲斐雅亮: "グアニン塩基に特異的なルミネッセンス試薬の活用法" 蛋白質核酸酵素. 42(4)(予定). (1997)

Masaaki Kai：“鸟嘌呤碱基特异性发光试剂的利用”蛋白质核酸酶 42(4)（计划）。