FLUORESCENT-LABELED OLIGONUCLEOTIDE PROBES - DETECTION OF HYBRID FORMATION IN SOLUTION BY FLUORESCENCE POLARIZATION SPECTROSCOPY
FLUORESCENT-LABELED OLIGONUCLEOTIDE PROBES - DETECTION OF HYBRID FORMATION IN SOLUTION BY FLUORESCENCE POLARIZATION SPECTROSCOPY
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DOI：
10.1093/nar/19.15.4097
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发表时间：
1991-08-11
影响因子：
14.9
通讯作者：
MAKINO, K
中科院分区：
文献类型：
--
作者：
MURAKAMI, A;NAKAURA, M;MAKINO, K
合成荧光素标记的寡核苷酸作为 DNA 探针，并用于监测杂交体的形成，即检测溶液中的 DNA 或寡核苷酸序列。将荧光素引入寡核苷酸是通过以乙二胺或六亚甲基二胺作为系链氧化膦酸氢键并随后用 FITC 标记伯胺部分来进行的。采用荧光各向异性 r 作为监测 F 探针在溶液中行为的指标。随着 F 探针链长的增加，观察到各向异性增加。当F-Probe与其互补寡核苷酸在溶液中形成杂合体时，与F-Probe本身相比，r值增加。这些观察结果清楚地表明，溶液中 r 的测量将很容易导致监测溶液中杂合体的存在。因此，使用荧光标记的寡核苷酸检测溶液中的某些核酸序列是有希望的。
Fluorescein-labeled oligonucleotides as DNA-probes were synthesized and used to monitor hybrid formation, namely to detect DNA or oligonucleotide sequence in solution. The introduction of fluorescein to oligonucleotides was carried out by oxidation of a hydrogen phosphonate linkage with ethylenediamine or hexamethylenediamine as a tether and by a subsequent labeling of the primary amine moiety by FITC. Fluorescence anisotropy, r, was adopted as an index to monitor the behavior of F-probe in solution. An increase in the anisotropy was observed upon an increase in the chain-length of F-probe. When F-Probe formed a hybrid with its complementary oligonucleotide in solution, the r value increased compared to that of F-Probe itself. These observations clearly indicate that measurements of r in solution will readily lead to the monitoring of the presence of a hybrid in solution. Consequently, it is promising to detect a certain nucleic acid sequence in solution using fluorescent-labeled oligonucleotides.