Sequence-specific fluorescence detection of DNA by polyamide-thiazole orange conjugates
Sequence-specific fluorescence detection of DNA by polyamide-thiazole orange conjugates
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DOI：
10.1021/ja054650k
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发表时间：
2005-11-30
影响因子：
15
通讯作者：
Dervan, PB
中科院分区：
文献类型：
--
作者：
Fechter, EJ;Olenyuk, B;Dervan, PB
不需要变性就能检测特异性双链DNA序列的荧光方法在遗传学领域可能是有用的。靶向其各自序列5 ′-WGGGWW-3 ′、5 ′-WGGCCW-3 ′和5 ′-WGWWCW-3 ′（W = A或T）的三种发夹吡咯-咪唑聚酰胺2-4在C-末端与噻唑橙子染料缀合，以在存在和不存在匹配双链体DNA的情况下检查其荧光性质。缀合物在不存在DNA的情况下发出弱荧光，但在加入1当量的匹配DNA时显示出显著增强（> 1000倍），而在加入错配DNA时仅显示出轻微增强。聚酰胺-染料缀合物以高亲和力（K-a > 10（8）M-1）结合特异性DNA序列，并通过嵌入（解旋角，Phi，类似于8度）解旋DNA双链体。这类新的聚酰胺提供了一种在不变性的情况下特异性检测DNA序列的方法。
Fluorescent methods to detect specific double-stranded DNA sequences without the need for denaturation may be useful in the field of genetics. Three hairpin pyrrole-imidazole polyamides 2-4 that target their respective sequences 5'-WGGGWW-3', 5'-WGGCCW-3', and 5'-WGWWCW-3'(W = A or T) were conjugated to thiazole orange dye at the C-termini to examine their fluorescence properties in the presence and absence of match duplex DNA. The conjugates fluoresce weakly in the absence of DNA but showed significant enhancement (> 1000-fold) upon the addition of 1 equiv of match DNA and only slight enhancement with the addition of mismatch DNA. The polyamide-dye conjugates bound specific DNA sequences with high affinity (K-a > 10(8) M-1) and unwound the DNA duplex through intercalation (unwinding angle, Phi, similar to 8 degrees). This new class of polyamides provides a method to specifically detect DNA sequences without denaturation.