天青Ⅰ分光光度法测定DNA

Spectrophotometric Determination of Deoxyribonucleic Acid Labeled with Azure Ⅰ

研究了生物染色剂天青 与脱氧核糖核酸在碱性条件下的结合反应 ,影响因素和最佳反应条件 ,建立了一种测定 DNA的新方法。在 p H=8.4的三羟甲基氨基甲烷 -盐酸 (Tris- HCl)缓冲溶液中 ,鱼精 DNA(fs DNA)及小牛胸腺 DNA(ct DNA)的线性范围分别为 0 .5— 10 .0 mg· L-1(fs DNA)、0 .5— 8.0 mg·L-1(ct DNA) ,相关系数为 0 .996 8(fs DNA)、0 .996 5 (ct DNA) ,表观摩尔吸光系数 ε60 9.90 nm(fs DNA) =2 .5×10 4L· mol-1· cm-1、ε60 9.90 nm(ct DNA) =3.1× 10 4L· mol-1· cm-1,用于合成样品的测定 。

A new spectrophotometric method for the determination of DNA was established based on the interaction between Azure Ⅰ and deoxyribonucleic acid(DNA) in pH8.4 medium.The effects of interference substances and optimization of procedure were investigated.The linear ranges are 0.5—10.0mg/L for fish sperm DNA and 0.5—8.0mg/L for calf thymus DNA.The correlation coefficient are 0.9968(fsDNA) and 0.9965(ctDNA),and the apparent molar absorption coefficient are ε 609.90nm (fsDNA)=2.5×10 4L·mol -1 ·cm -1 、 ε 609.90nm (ctDNA)=3.1×10 4L·mol -1 ·cm -1 ,respectively.The method has been applied to the determination of DNA in mixed samples with satisfactory results.