应用多重标记引物增强寡核苷酸芯片的荧光信号强度(英文)

Multiply Labeled Primers Amplifying Fluorescent Signal on Oligonucleotide Microarray

寡核苷酸芯片技术是一种高通量发掘和采集生物信息的强大技术平台,目前已广泛应用于生物科学领域.为改善寡核苷酸芯片的分析性能,对影响芯片杂交结果的因素,如片基表面的化学处理、探针的长度、间隔臂的长度、杂交条件等,进行了深入的研究和优化.对寡核苷酸芯片而言,仍有待解决的问题是如何产生更强的荧光信号来改善其检测灵敏度.利用两种类型的多个荧光分子标记的引物,来增强二维寡核苷酸芯片平面上的荧光信号强度.两种引物分别命名为:多标记线性引物和多标记分支引物.通过增加标记在目标DNA 片段上的荧光分子数,可以显著增强寡核苷酸芯片上相应捕获探针的信号强度.实验表明,使用多标记引物能将所用的寡核苷酸微阵列的检测限(以能够检测的最低模板量计算)降低至单荧光标记引物的1/100 以下,多重标记技术是一种有效增强微型化探针矩阵检测灵敏度的信号放大方法.

Oligonucleotide microarray technology is a powerful data-mining platform and has been widely applied in biosciences. To improve the performance of assays on the oligonucleotide microarray, the factors that influence the hybridization effects such as surface chemistry, probe size, spacer length, hybridization conditions etc were intensely studied and optimized. However, it is a key problem with DNA microarrays how to generate higher fluorescent signals to improve the detection sensitivity. Two types of multiply labeled primers, termed multiply labeled linear primer and multiply labeled branched primer, were used to enhance the fluorescent signal obtained from two-dimensional DNA microarrays. The signal was intensified by increasing the number of fluorophores labeled on the target DNA segment. It was indicated that the detection limit （minimum template amount for detection） of the multiply labeled primers is about 1% of that of the singly labeled primer. Multiple labeling is an effective signal amplification method to increase the detection sensitivity of the probes in a miniaturized array format.