Use of oligonucleotides with modified bases in hybridization of nucleic acids
A technology of oligonucleotide and nucleic acid, which is applied in the application field of oligonucleotides with modified bases in nucleic acid hybridization, and can solve problems such as differences in stability
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[0093] 讲述了上述含磷连键的制备的代表性美国专利包括但不限于U.S.:3,687,808,4,469,863,4,476,301,5,023,243,5,177,196,5,188,897,5,264,423,5,276,019,5,278,302,5,286,717,5,321,131,5,399,676,5,405,939,5,453,496,5,455,233, 5,466,677,5,476,925,5,519,126,5,536,821,5,541,306,5,550,111,5,563,253,5,571,799,5,587,361,5,194,599,5,565,555,5,527,899,5,721,218,5,672,697和5,625,050,它们每个在此引为参考。
[0094] Contemplated modified oligonucleotide backbones containing no phosphorus atoms therein, having short chain alkyl or cycloalkyl internucleoside linkages, mixed heteroatoms and alkyl or cycloalkyl internucleoside linkages, Or one or more short-chain heteroatoms or a backbone formed by linkages between heterocyclic nucleosides. They include backbones with morpholino linkages (partially formed from the sugar moieties of nucleosides); siloxane backbones; thioether, sulfoxide, and sulfone backbones; formyl and thioformyl backbones; methyleneformyl and Thioformyl skeleton; riboseacetyl skeleton; skeleton containing alkenes; sulfa...
Embodiment 1
[0147] Modified / unmodified antisense oligonucleotides with complementary membrane-bound unmodified sense oligonucleotides Nucleotide binding
[0148]Each of the following patent applications is incorporated by reference in its entirety: U.S. Serial No. 60 / 797,448, filed May 3, 2006, and 11 / 742,384, filed April 30, 2007 (referred to as U.S. Patent Application Publication No. US 2007 / 0259830 published).
[0149] To compare the relative binding efficiency of oligonucleotides containing one or more modified oligonucleotides, modified oligonucleotides are generated and compared to the hybridization efficiency of unmodified oligonucleotides. Binding efficiency is determined relative to the pH of the hybridization reaction.
[0150] Oligonucleotides contain a modified base 5-hydroxycytosine (C * ) and 8-hydroxyguanine (G * ) or both. Unmodified oligonucleotides were used as controls. Oligonucleotides are shown in Table 1.
[0151] Table 1
[0152] Oligonucleotide...
Embodiment 2
[0167] Example 2, Modified / unmodified antisense oligonucleotides and complementary membrane-bound DNA combine
[0168] Similar experiments were performed using the same method to measure the relative binding efficiency of oligonucleotides to DNA. Still using a 4th order polynomial to fit the data for each modified oligonucleotide, the corresponding plots are in image 3 given in.
[0169] In contrast to oligonucleotide-oligonucleotide binding, in this case the pH dependence has three maxima, but the highest is still at lower pH values (approximately pH=4.5...5.0). This maximum is also significantly higher than oligonucleotide-oligonucleotide binding. The relative binding efficiencies were 2.9 and 2.1 for modified oligonucleotides f2 and f1, respectively. The other two maxima of the pH-dependent relative binding efficiencies of the modified oligonucleotides were located at approximately pH=7.5 and pH=10 ( image 3 ).
[0170] Apparently, oligonucleotide f with two mo...
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