Synthesis And Covalent Attachment Of A Methylene Blue Derivative To A Triple Helix Forming Oligonucleotide
| Main Author: | Purwanto, Maria Goretti Marianti |
|---|---|
| Format: | Article PeerReviewed application/pdf |
| Terbitan: |
Research India Publications
, 2015
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| Subjects: | |
| Online Access: |
http://repository.ubaya.ac.id/26574/1/Purwanto_Synthesis%20and%20Covalent_2015.pdf http://www.ripublication.com http://repository.ubaya.ac.id/26574/ |
Daftar Isi:
- Photoactive methylene blue (MB) is a cationic planar chromophore which is able to interact with double and triple helical DNA. Thus on its applications can be used in the development of a new drug that specifically target a DNA sequence. Covalent attachment of the MB to the DNA through a flexible heptamethylene linker has been accomplished, enabling the defined positioning of the dye moiety at specific sites of the DNA strands by forming a triple helix. The structure of MB itself does not provide a suitable functional group for a direct attachment to an oligonucleotide. Thus, the MB derivative, 3 - N - (4 - carboxybutyl) - methylamino - 7 - dimethylaminophenaza - thionum chloride was synthesized. The attachment of the MB derivative to a 3’ - aminoalkylated oligonucleotide was accomplished by the activation of the carboxyl group with a yield from 40% - 70%. A precipitation step using n - butanol was performed before HPLC purification in order to remove the excess of activated MB ester. Unreacted oligonucleotide was identified in the chromatogram from its absorbance at the typical wavelength for nucleic acids and its lack of absorbance at 660 nm. The conjugate products bearing the MB moiety were recognized from the additional absorbance at around 660 nm. Another confirmation comes from the MS data. 1D 1H NMR spectra of the MB - oligonucleotide conjugates as well as the non - conjugated 3’ - aminoalkyl - oligonucleotide in H2O are acquired to compare the oligonucleotide before and after conjugation with MB.