快速简便的杂交检测DNA(英文)

A Simple Solution Hybridization Rapidly Detects DNA by Fluorescent Probe

Southern杂交作为黄金标准,已广泛运用于DNA的检测上。但是,经典的southern杂交和近年来一些DNA检测方法存在放射性污染,操作繁琐,耗时,对实验仪器设备要求高等问题。本文利用异硫氰酸荧光素(FITC)标记的dUTP合成DNA探针建立了快速检测DNA的液相杂交方法。该方法包括探针制备、液相杂交、电泳分离和信号检测四个步骤,并在此基础上对FITC-双链和FITC-单链探针的杂交效果作了比较。结果显示,使用FITC标记的两种探针都能取得良好的实验结果,但单链探针较双链的检测灵敏度高;双链DNA探针可以检测出0.8μg(3.64×10-13 mol)的质粒,而单链DNA探针可以检测出0.38μg(1.82×10-13 mol)的质粒DNA,在检出效率上是前者的2.1倍。整个检测过程操作简便,可在3h内完成,可较好地解决了其他DNA检测方法存在的费时费力的问题。

Traditional Southern blot,as the golden standard to detect DNA,is widely used in molecular biolo-gy.However,its operation process has some significant disadvantages,such as radioactive contamination, complex procedures,time-consuming.Some new methods developed in recent years need expensive equipments and also have the defects of complex procedures and time-consuming.The paper described a method to detect DNA quickly by liquid hybridization.The probe used is a fragment of DNA sequence labeled by FITC-12-dUTP.The whole process includes four steps：probe preparation,hybridization,electrophoresis and signal de-tection.The comparative experiments using the single strand DNA probe and the double strand DNA probe in-dicated that the sensitivity of single strand DNA probes,which could detect 0. 38μg （1. 82 ×10^-13 mol）plas-mid,is 2. 1 times than that of the double strand DNA probes which could detect 0. 8μg（3. 64×10^-13 mol） plasmid.The total procedure is simple and can be completed in 3 h.